Non-benzenoid aromatic systems for imaging, monitoring and therapy

ABSTRACT

This invention is directed to non-benzenoid aromatic compounds. Other aspects include methods of using non-benzenoid aromatic compounds for imaging and phototherapeutic uses thereof. Non-benzenoid compounds provided herein generally have one or more substituent groups which allow tailoring of the spectral properties or provide photoreactivity or targeting ability.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.13/395,493, filed Mar. 30, 2012, which itself is a 371 national phasefiling of PCT application no. PCT/US2010/048347, filed Sep. 10, 2010,which claims the benefit of priority of U.S. Provisional ApplicationsNo. 61/241,624, filed Sep. 11, 2009, the disclosures of which are herebyincorporated by reference as if written herein in their entireties.

INCORPORATION OF SEQUENCE LISTING

A copy of the Sequence Listing and a computer readable form of thesequence containing the file named “33693-192_Seq_Listing_ST25.txt”,which is 582 bytes in size (as measured in MICROSOFT WINDOWS® EXPLORER),are provided herein and are herein incorporated by reference. ThisSequence Listing consists of SEQ ID NO:1.

BACKGROUND

Optical agents currently play a central role in a large number of invivo, in vitro and ex vivo clinical procedures including importantdiagnostic and therapeutic procedures. Photodiagnostic andphototherapeutic agents, for example, include a class of moleculescapable of absorbing, emitting, and/or scattering electromagneticradiation applied to a biological material, particularly in the visibleand/or near infrared regions of the electromagnetic spectrum. Thisproperty of optical agents is used in a range of biomedical applicationsfor visualizing, imaging or otherwise characterizing biologicalmaterials and/or achieving a desired therapeutic outcome. Recentdevelopments in targeted administration and delivery of optical agents,and advanced systems and methods for applying and detectingelectromagnetic radiation in biological environments has considerablyexpanded the applicability and effectiveness of optical agents forclinical applications.

Important applications of optical agents that absorb and/or emit in thevisible and/or near-infrared (NIR) region of the electromagneticspectrum include their use in biomedical imaging and visualization. Forexample, compounds absorbing and/or emitting light in these regions ofthe electromagnetic spectrum currently are useful for opticaltomography, optoacoustic tomography, optical coherence tomography,confocal scanning laser tomography, optical coherence tomography, andfluorescence endoscopy; techniques which have emerged as essentialmolecular imaging techniques for imaging and visualizing biologicalprocesses at the organ, cellular and subcellular (e.g., molecular)levels. Biomedical images are generated, for example, by detectingelectromagnetic radiation, nuclear radiation, acoustic waves, electricalfields, and/or magnetic fields transmitted, emitted and/or scattered bycomponents of a biological sample. Modulation of the energy or intensityof the applied radiation yields patterns of transmitted, scatteredand/or emitted radiation, acoustic waves, electrical fields or magneticfields that contain useful anatomical, physiological, and/or biochemicalinformation. A number of applications of biomedical imaging have maturedinto robust, widely used clinical techniques including planar projectionand tomographic X-ray imaging, magnetic resonance imaging, ultrasoundimaging, and gamma ray imaging.

Established optical imaging and visualization techniques are based onmonitoring spatial variations in a variety of optical parametersincluding the intensities, polarization states, and frequencies oftransmitted, reflected, and emitted electromagnetic radiation. Giventhat many biological materials of interest are incompatible withultraviolet light, research is currently directed to developing andenhancing imaging techniques using visible and near infrared (NIR)radiation (from about 400 nm to about 900 nm). In particular, NIR light(700 nm to 900 nm) is useful for visualizing and imaging deeper regionsthan visible light because electromagnetic radiation of this wavelengthrange is capable of substantial penetration (e.g., up to fourcentimeters) in a range of biological media. Optical imaging andvisualization using optical agents has potential to provide a lessinvasive and safer imaging technology, as compared to X-ray, and otherwidely used nuclear medicine technologies. Applications of opticalimaging for diagnosis and monitoring of the onset, progression andtreatment of various disease conditions, including cancer, are wellestablished. (See, e.g., D. A. Benaron and D. K. Stevenson, Opticaltime-of-flight and absorbance imaging of biologic media, Science, 1993,259, pp. 1463-1466; R. F. Potter (Series Editor), Medical opticaltomography: functional imaging and monitoring, SPIE Optical EngineeringPress, Bellingham, 1993; G. J. Tearney et al., In vivo endoscopicoptical biopsy with optical coherence tomography, Science, 1997, 276,pp. 2037-2039; B. J. Tromberg et al., Non-invasive measurements ofbreast tissue optical properties using frequency-domain photonmigration, Phil. Trans. Royal Society London B, 1997, 352, pp. 661-668;S. Fantini et al., Assessment of the size, position, and opticalproperties of breast tumors in vivo by noninvasive optical methods,Appl. Opt., 1998, 37, pp. 1982-1989; A. Pelegrin et al.,Photoimmunodiagnosis with antibody-fluorescein conjugates: in vitro andin vivo preclinical studies, J. Cell Pharmacol., 1992, 3, pp. 141-145).

Optical agents for in vivo and in vitro biomedical imaging, anatomicalvisualization and monitoring organ function are described inInternational Patent Publication WO2008/108941; U.S. Pat. Nos.5,672,333; 5,698,397; 6,167,297; 6,228,344; 6,748,259; 6,838,074;7,011,817; 7,128,896, and 7,201,892. In this context, optical imagingagents are commonly used for enhancing signal-to-noise and resolution ofoptical images and extending these techniques to a wider range ofbiological settings and media. In addition, use of optical imagingagents having specific molecular recognition and/or tissue targetingfunctionality has also been demonstrated as effective for identifying,differentiating and characterizing discrete components of a biologicalsample at the organ, tissue, cellular, and molecular levels. Further,optical agents have been developed as tracers for real time monitoringof physiological function in a patient, including fluorescence-basedmonitoring of renal function. (See International Patent PublicationPCT/US2007/0149478). Given their recognized utility, considerableresearch continues to be directed toward developing improved opticalagents for biomedical imaging and visualization.

In addition to their important role in biomedical imaging andvisualization, optical agents capable of absorption in the visible andNIR regions have also been extensively developed for clinicalapplications for phototherapy. The benefits of phototherapy usingoptical agents are widely acknowledged as this technique has thepotential to provide efficacy comparable to radiotherapy, while entirelyavoiding exposure of non-target organs and tissue to harmful ionizingradiation. Photodynamic therapy (PDT), in particular, has been usedeffectively for localized superficial or endoluminal malignant andpremalignant conditions. The clinical efficacy of PDT has also beendemonstrated for the treatment of various other diseases, injuries, anddisorders, including cardiovascular disorders such as atherosclerosisand vascular restenosis, inflammatory diseases, ophthalmic diseases anddermatological diseases. Visudyne and Photofrin, for example, are twooptical agents that have been developed for the treatment of maculardegeneration of the eye and for ablation of several types of tumors,respectively. (See, e.g., Schmidt-Drfurth, U.; Bringruber, R.; Hasan, T.Phototherapy in ocular vascular disease. IEEE Journal of Selected Topicsin Quantum Electronics 1996, 2, 988-996; Mlkvy, P.; Messmann, H.;Regula, J.; Conio, M.; Pauer, M.; Millson, C. E.; MacRobert, A. J.;Brown, S. G. Phototherapy for gastrointestinal tumors using threephotosensitizers—ALA induced PPIX, Photofrin, and MTHPC. A pilot study.Neoplasma 1998, 45, 157-161; Grosjean, P.; Wagieres, G.; Fontolliet, C.;Van Den Bergh, H.; Monnier, P. Clinical phototherapy for superficialcancer in the esophagus and the bronchi: 514 nm compared with 630 nmlight irradiation after sensitization with Photofrin II. British Journalof Cancer 1998, 77, 1989-1955; Mitton, D.; Ackroyd, R. Phototherapy ofBarrett's oesophagus and oesophageal carcinoma—how I do it.Photodiagnostics and Phototherapy 2006, 3, 96-98; and Li, L.; Luo, R.;Liao, W.; Zhang, M.; Luo, Y.; Miao, J. Clinical study of photofrinphototherapy for the treatment of relapse nasopharyngeal carcinoma.Photodiagnostics and Phototherapy 2006, 3, 266-271; See, Zheng Huang “AReview of Progress in Clinical Photodynamic Therapy”, Technol Cancer ResTreat. 2005 June; 4(3): 283-293; “Photodiagnosis and PhotodynamicTherapy”, Brown S, Brown E A, Walker I. The present and future role ofphotodynamic therapy in cancer treatment. Lancet Oncol. 2004; 5:497-508;Triesscheijn M, Baas P, Schellens J H M. “Photodynamic Therapy inOncology”; The Oncologist. 2006; 11:1034-1044; and Dougherty T J, GomerC J, Henderson B W, Jori G, Kessel D, Korbelik M, Moan J, Peng Q.Photodynamic Therapy. J. Natl. Cancer Inst. 1998; 90:899-905).

Phototherapy is carried out by administration and delivery of aphotosensitizer to a therapeutic target tissue (e.g., tumor, lesion,organ, etc.) followed by photoactivation of the photosensitizer byexposure to applied electromagnetic radiation. Phototherapeuticprocedures require photosensitizers that are relatively chemicallyinert, and become activated only upon irradiation with light of anappropriate wavelength. Selective tissue injury can be induced withlight when photosensitizers bind to the target tissues, either directlyor through attachment to a bioactive carrier or targeting moiety.Photosensitizers essentially operate via two different pathways,classified as Types 1 and 2. A primary distinction between these classesof photosensitizers is that the Type 1 process operates via directenergy or electron transfer from the photosensitizer to the cellularcomponents thereby inducing cell death, whereas the Type 2 processinvolves first the conversion of singlet oxygen from the triplet oxygenfound in the cellular environment followed by either direct reaction ofsinglet oxygen with the cellular components or further generatingsecondary reactive species (e.g. peroxides, hydroxyl radical, etc.)which will induce cell death.

The Type 1 mechanism proceeds via a multistep process involvingactivation of the photosensitizer by absorption of electromagneticradiation followed by direct interaction of the activatedphotosensitizer, or reactive intermediates derived from thephotosensitizer, with the target tissue, for example via energytransfer, electron transfer or reaction with reactive species (e.g.,radicals, ions, nitrene, carbene, etc.) resulting in tissue damage. TheType 1 mechanism can be schematically represented by the followingsequence of reactions:

wherein hv indicates applied electromagnetic radiation and(PHOTOSENSITIZER)* indicates excited state of the photosensitizer. TheType 2 mechanism proceeds via a three-step process involving activationof the photosensitizer by absorption of electromagnetic radiationfollowed by energy transfer from the activated photosensitizer to oxygenmolecules in the environment of the target tissue. This energy transferprocess generates excited state oxygen (¹O₂) which subsequentlyinteracts with the target tissue so as to cause tissue damage. The Type2 mechanism can be schematically represented by the following sequenceof reactions:

wherein hv indicates applied electromagnetic radiation,(PHOTOSENSITIZER)* indicates photoactivated photosensitizer, ³O₂ isground state triplet oxygen, and ¹O₂ is excited state singlet oxygen.

The biological basis of tissue injury brought about by tumorphototherapeutic agents has been the subject of intensive study. Variousbiochemical mechanisms for tissue damage have been postulated, whichinclude the following: a) cancer cells up-regulate the expression of lowdensity lipoprotein (LDL) receptors, and phototherapy (PDT) agents bindto LDL and albumin selectively; (b) porphyrin-like substances areselectively taken up by proliferative neovasculature; (c) tumors oftencontain increased number of lipid bodies and are thus able to bind tohydrophobic photosensitizers; (d) a combination of “leaky” tumorvasculature and reduced lymphatic drainage causes porphyrin accumulationreferred to as “EPR” (enhanced permeability and retention) effect; (e)tumor cells may have increased capabilities for phagocytosis orpinocytosis of porphyrin aggregates; (f) tumor associated macrophagesmay be largely responsible for the concentration of photosensitizers intumors; and (g) cancer cells may undergo apoptosis induced byphotosensitizers. Among these mechanisms, (f) and (g) are the mostgeneral and, of these two alternatives, there is a general consensusthat (f) is the most likely mechanism by which the phototherapeuticeffect of porphyrin-like compounds is induced.

Much of the research in the past several decades has focused ondeveloping phototherapeutic agents based on the Type 2 (PDT) mechanism.Surprisingly, there has been considerably less attention devoted to Type1 phototherapeutic agents despite the fact that there are numerousclasses of compounds that could potentially be useful for phototherapythat function via this mechanism. Unlike Type 2, the Type 1 process doesnot require oxygen; and hence Type 1 photosensitizers are expected to bepotentially more effective than Type 2 photosensitizers under hypoxicenvironments typically found in solid tumors. Second, the Type 1mechanism involves two steps (photoexcitation and direct energytransfer), whereas the Type 2 mechanism involves three steps(photoexcitation, singlet oxygen generation, and energy transfer).Further, studies have recently shown that production of high levels ofreactive oxygen species can induce an anti-inflammatory response, whichmay result in blood vessels to become more “leaky,” thereby increasingthe risk of metastasis (Chen, B.; Pogue, B.; Luna, J. M.; Hardman, R.L.; Hoopes, P. J.; Hasan, T. Tumor vascular permeabilization byvascular-targeting photosensitization: effects, mechanism, andtherapeutic implications. Clinical Cancer Research 2006, 12(3, Pt. 1),917-923). Targeted Type 1 photosensitizers, by their very nature, arenot expected to produce reactive oxygen species; rather, the reactivespecies produced by these photosensitizers will immediately react withthe cellular component at the binding site and trigger cell death. Type2 phototherapeutic agents, however, do have certain advantages over Type1 agents. For example, Type 2 agents can potentially be catalytic, i.e.,the Type 2 photosensitizer is regenerated once the energy transfer tothe oxygen has taken place. In contrast, Type 1 process would generallybe expected to require stoichiometric amounts of the photosensitizer insome clinical settings. Table I provides a summary of the attributes ofType 1 and Type 2 phototherapeutic agents. Given these attributes, it isclear that development of safe and effective Type 1 phototherapeuticagents would be useful to complement the existing therapeutic approachesprovided by Type 2 agents, and to enhance the therapeutic portfolioavailable for clinicians.

TABLE 1 Comparison between Type 1 and Type 2 processes for phototherapy.TYPE 1 PROCESS TYPE 2 PROCESS Two-step process. Three-step process. Notwell explored. Very well studied. Light of any wavelength Requires redlight for optimal can be used. performance. Does not require oxygen.Requires oxygen. Large classes of compounds. Limited classes ofcompounds. Stoichiometric. Potentially catalytic. Intramolecular energytransfer to Intermolecular energy transfer to generate reactive species.generate reactive oxygen species. No products in the market. Twoproducts are in use.

Specific optical, chemical and pharmacokinetic properties of opticalagents are necessary for their effective use in Type 1 and Type 2phototherapeutic applications. For example, optical agents for theseapplications preferably have strong absorption in the visible or NIRregions, and also exhibit low systemic toxicity, low mutagenicity, andrapid clearance from the blood stream. These optical agents must also becompatible with effective administration and delivery to the targettissue, for example by having reasonable solubilities and a low tendencyfor aggregation in solution. Upon excitation by absorption of visibleand NIR electromagnetic radiation, optical agents for Type 1 and 2phototherapy preferably provide large yields of singlet oxygen (Type 2)or other reactive species, such as free radicals or ions, capable ofcausing local tissue damage. Both Type 1 and Type 2 photosensitizerstypically undergo photoactivation followed by intersystem crossing totheir lowest triplet excited state, and therefore, a relatively longtriplet lifetime is usually beneficial for providing effective tissuedamage. Other useful properties of optical agents for these applicationsinclude chemical inertness and stability, insensitivity of opticalproperties to changes in pH, and compatibility with conjugation toligands providing targeted delivery via molecular recognitionfunctionality. Multifunctional optical agents have also been developedfor phototherapy that are capable of providing both imaging and visualfunctionality upon excitation at a first range of wavelengths andphototherapeutic functionality upon excitation at a second range ofwavelength. (See, U.S. Pat. No. 7,235,685 and International PatentPublication WO 2007/106436).

Optical agents for phototherapeutic applications preferably exhibit ahigh degree of selectivity for the target tissue. Selectivity providedby optical agents facilitates effective delivery to a target tissue ofinterest and provides a means of differentiating different tissueclasses during therapy. Selective tissue injury can be induced withlight when photosensitizers bind to the target tissues either directly,as in the case of Photofrin, or through attachment to a bioactivecarrier, or through in situ biochemical synthesis of the photosensitizerin localized area, as in the case of 2-aminolevulinic acid, which is anintermediate in the biosynthesis of porphyrin. Previous studies haveshown that certain dyes localize in tumors and serve as a powerful probefor the detection and treatment of small cancers. (D. A. Belinier etal., Murine pharmacokinetics and antitumor efficacy of the photodynamicsensitizer 2[I-hexyloxyethyl]-2-devinyl pyropheophorbide-a, J.Photochem. Photobiol., 1993, 20, pp. 55-61; G. A. Wagnieres et al., Invivo fluorescence spectroscopy and imaging for oncological applications,Photochem. Photobiol., 1998, 68, pp. 603-632; J. S. Reynolds et al.,Imaging of spontaneous canine mammary tumors using fluorescent contrastagents, Photochem. Photobiol., 1999, 70, pp. 87-94). It is generallyrecognized, however, that many of these dyes do not localizepreferentially in malignant tissues. A number of strategies have beendeveloped for imparting selectivity and/or targeting functionality byincorporation of a molecular recognition component in the optical agent.For example, targeting of fluorescent dyes to tumors has beendemonstrated by us and others using dye conjugates with antibodies andpeptides for diagnostic imaging of tumors. (See, Achilefu et al., Novelreceptor-targeted fluorescent contrast agents for in vivo imaging oftumors, Investigative Radiology, 2000, 35, pp. 479-485; Ballou et al.,Tumor labeling in vivo using cyanine conjugated monoclonal antibodies,Cancer Immunology and Immunotherapy, 1995, 41, pp. 257-263; and Licha etal., New contrast agent for optical imaging: acid cleavable conjugatesof cyanine dyes with biomolecules, in Biomedical Imaging: Reporters,Dyes and Instrumentation, Proceedings of SPIE, 1999, 3600, pp. 29-35).Therefore, receptor-target mediated phototherapy agents provide apromising pathway for achieving site selective activation at varioustarget tissues.

For both photodiagnostic and phototherapeutic applications, opticalagents preferably exhibit a high degree of selectivity for the targettissue. Selectivity provided by optical agents facilitates effectivedelivery to a target tissue of interest and provides a means ofdifferentiating different tissue classes during imaging, visualizationand therapy. There is a considerable need for developing optical agentsfor biomedical applications that have high absorption/emissionproperties in the visible and NIR regions, high photostability,insensitivity to pH, and wavelength tunability, as well as selectivityfor the target tissue.

SUMMARY

The invention relates generally to optical agents for biomedicalapplications, including imaging, diagnosing and/or treating medicalconditions. Compounds provided absorb and emit spectral energy in thevisible, near infrared, and/or any other wavelength range useful foroptical detection in medical procedures. The invention includes opticalagents and related therapeutic methods, comprising non-benzenoidaromatic compounds having substituent groups which allow tailoring ofthe spectral properties and/or provide photoreactivity and/or targetingability.

In embodiments, compounds of the invention have the formula (FX1):

wherein:

X_(a) is N or —C-(L⁶)_(q)-W⁶—R⁶;

X_(b) is N or —C-(L⁷)_(q)-W⁷—R⁷;

X_(c) is N or —C-(L⁸)_(q)-W⁸—R⁸;

X_(d) is N or —C-(L¹)_(q)-W¹—R¹;

X_(e) is N or —C-(L²)_(q)-W²—R²;

each of L¹ to L⁸ is independently C₁-C₁₀ alkylene, C₃-C₁₀ cycloalkylene,C₂-C₁₀ alkenylene, C₃-C₁₀ cycloalkenylene, C₂-C₁₀ alkynylene,ethenylene, ethynylene, phenylene, 1-aza-2,5-dioxocyclopentylene,—(CH₂CH₂O)_(m)—, —(CHOH)_(m)—, or 1,4-diazacyclohexylene;

each m is independently an integer selected from the range of 1 to 100;

each q is independently 0 or 1;

each of W¹ to W⁸ is independently a single bond, —(CH₂)_(n)—,—O(CH₂)_(n)—, —(CH₂)_(n)O—, —(HCCH)_(n)—, —O—, —S—, —SO—, —SO₂—, —SO₃ ⁻,—OSO₂—, —NR⁹—, —CO—, —COO—, —OCO—, —OCOO—, —CONR¹⁰—, —NR¹¹CO—,—OCONR¹²—, —NR¹³COO—, —NR¹⁴CONR¹⁵—, —NR¹⁶CSNR¹⁷—, —O(CH₂)_(n)—,—S(CH₂)_(n)—, —NR¹⁸(CH₂)_(n)—, —CO(CH₂)_(n)—, —COO(CH₂)_(n)—,—OCO(CH₂)_(n)—, —OCOO(CH₂)_(n)—, —CONR¹⁹(CH₂)_(n)—,—CONR²⁰(CH₂)_(n)(OCH₂CH₂)_(u)—, —NR²¹CO(CH₂)_(n)—, —OCONR²²(CH₂)_(n)—,—NR²³COO(CH₂)_(n)—, —NR²⁴CONR²⁵(CH₂)_(n)—, —NR²⁶CSNR²⁷(CH₂)_(n)—,—O(CH₂)_(n)NR²⁸CO(CH₂)_(n)—,—CO(CH₂)_(n)(CH₂OCH₂)(CH₂)_(n)NR²⁹(CH₂)_(n)NR³⁰CO—, —NR⁶⁹SR⁷⁰—, or—CO(CH₂)_(n)NR³¹CO—;

each n is independently an integer selected from the range of 1 to 10;

each of R⁹ to R³¹ and each of R⁶⁹ to R⁷⁰ is independently hydrogen,C₁-C₂₀ alkyl, or C₅-C₃₀ aryl;

each of R¹ to R⁸ is independently hydrogen, C₁-C₂₀ alkyl, C₅-C₃₀ aryl,C₁-C₂₀ acyl, C₂-C₂₀ alkenyl, C₂-C₂₀ alkynyl, C₅-C₂₀ alkylaryl, C₁-C₆alkoxycarbonyl, halo, halomethyl, dihalomethyl, trihalomethyl, —CN,—CO₂R³², —OCONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷, —SO₂R³⁸, —SO₂OR³⁹,—SO₂NR⁴⁰R⁴¹, —PO₃R⁴²R⁴³, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, —NR⁴⁸COR⁴⁹,—CH₂(CHOH)_(n)R⁵, —(CH₂CH₂O)_(u)R⁵¹, —CH(R⁵²)CO₂H, —CH(R⁵³)NH₂, TG¹ toTG⁸, PS¹ to PS⁸, or FL¹ to FL⁸;

each u is independently an integer selected from the range of 1 to 25;

each of R³² to R⁵⁵ is independently hydrogen or C₁-C₁₀ alkyl;

each of TG¹ to TG⁸ is independently an amino acid, a peptide, a protein,a nucleoside, a nucleotide, an enzyme, a carbohydrate, a glycomimetic,an oligomer, a lipid, a polymer, an antibody, an antibody fragment, amono- or polysaccharide comprising 1 to 50 carbohydrate units, aglycopeptide, a glycoprotein, a peptidomimetic, a drug, a drug mimic, ahormone, a receptor, a metal chelating agent, a radioactive ornonradioactive metal complex, a mono- or polynucleotide comprising 1 to50 nucleic acid units, a polypeptide comprising 2 to 30 amino acidunits, or an echogenic agent;

each of PS¹ to PS⁸ is independently a photosensitizing moiety capable ofproducing one or more free radicals, nitrenes, carbenes, and/or singletoxygen, and wherein each of PS¹ to PS⁸ comprises at least one azide,azo, diazo, oxaza, diaza, dithia, thioxa, dioxa, phthalocyanine,rhodamine, or porphyrin group; and each of FL¹ to FL⁸ is independently afluorescent moiety selected from naphthoquinones, anthracenes,anthraquinones, phenanthrenes, tetracenes, naphthacenediones, pyridines,quinolines, isoquinolines, indoles, isoindoles, pyrroles, imidiazoles,pyrazoles, pyrazines, purines, benzimidazoles, benzofurans,dibenzofurans, carbazoles, acridines, acridones, phenanthridines,thiophenes, benzothiophenes, dibenzothiophenes, xanthenes, xanthones,flavones, coumarins, phenoxazines, phenothiazines, phenoselenazines,cyanines, indocyanines, and azo compounds; wherein any adjacent R¹ to R⁸may combine, optionally with one or two —CR⁵⁴R⁵⁵ groups, to form C₃-C₇cycloalkyl, C₃-C₇ heterocycloalkyl, C₆ aryl, or C₅-C₆ heteroaryl;

wherein at least one of R¹ to R⁸ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵,—NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹; and wherein at least one of R¹ to R⁸ is halo,trihalomethyl, —CN, —CO₂R³², —OCONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷,—SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₁-C₁₀ acyl; or wherein at least one of-(L¹)_(q)-W¹—R¹, -(L²)_(q)-W²—R², -(L³)_(q)-W³—R³, -(L⁴)_(q)-W⁴—R⁴,-(L⁵)_(q)-W⁵—R⁵, -(L⁶)_(q)-W⁶—R⁶, -(L⁷)_(q)-W⁷—R⁷, or -(L⁸)_(q)-W⁸—R⁸includes —(OCH₂CH₂)_(u)—.

In an embodiment, at least one of R¹ to R⁸ is independently C₁-C₆ alkyl,—OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹. In an embodiment, at least one ofR¹ to R⁸ is independently halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, or —SO₂NR⁴⁰R⁴¹. In an embodiment, atleast one of R¹ to R⁸ is independently a targeting group TG¹ to TG⁸. Inan embodiment, at least one of R¹ to R⁸ is independently aphotosensitizer PS¹ to PS⁸. In an embodiment, at least one of R¹ to R⁸is independently a fluorophore FL¹ to FL⁸. In an embodiment, at leastone PS¹ to PS⁸ comprises an azide, azo, diazo, oxaza, diaza, thioxa,phthalocyanine, rhodamine or porphyrin group.

In an embodiment, at least one of -(L¹)_(q)-W¹—R¹, -(L²)_(q)-W²—R²,-(L³)_(q)-W³—R³, -(L⁴)_(q)-W⁴—R⁴, -(L⁵)_(q)-W⁵—R⁵, -(L⁶)_(q)-W⁶—R⁶,-(L⁷)_(q)-W⁷—R⁷, or -(L⁸)_(q)-W⁸—R⁸ includes a —(OCH₂CH₂)_(u)— group.

In the embodiment where any adjacent R¹ to R⁸ may combine, optionallywith one or two —CR⁵⁴R⁵⁵ groups, to form C₃-C₇ cycloalkyl, C₃-C₇heterocycloalkyl, C₆ aryl, or C₅-C₆ heteroaryl, the C₃-C₇ cycloalkyl,C₃-C₇ heterocycloalkyl, C₆ aryl, or C₅-C₆ heteroaryl groups may beoptionally substituted with one or more substituents. The optionalsubstituents may include, for example, one or more of C₁-C₂₀ alkyl,C₅-C₃₀ aryl, C₁-C₂₀ acyl, C₂-C₂₀ alkenyl, C₂-C₂₀ alkynyl, C₅-C₂₀alkylaryl, C₁-C₆ alkoxycarbonyl, halo, halomethyl, dihalomethyl,trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷,—SO₂R³⁸, —SO₂OR³⁹, —SO₂NR⁴⁰R⁴¹, —PO₃R⁴²R⁴³, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷,—NR⁴⁸COR⁴⁹, —CH₂(CHOH)_(n)R⁵⁰, —(CH₂CH₂O)_(u)R⁵¹, —CH(R⁵²)CO₂H,—CH(R⁵³)NH₂, TG¹ to TG⁸, PS¹ to PS⁸, or FL¹ to FL⁸, where each of R³² toR⁵⁵ is independently hydrogen or C₁-C₁₀ alkyl and TG¹ to TG⁸, PS¹ toPS⁸, and FL¹ to FL⁸ are each as defined above for Formula (FX1). In anembodiment where any adjacent R¹ to R⁸ may combine, optionally with oneor two —CR⁵⁴R⁵⁵ groups, to form C₃-C₇ heterocycloalkyl, or heteroaryl,the C₃-C₇ heterocycloalkyl, or C₅-C₆ heteroaryl groups may contain oneor more heteroatoms which may be the same or different.

In an embodiment, the invention provides compounds having formula (FX2)to (FX6) where the variables are as defined above for formula (FX1):

wherein X is N or —C-(L⁸)_(q)-W⁸—R⁸;

wherein X is N or —C-(L⁷)_(q)-W⁷—R⁷;

wherein X is N or —C-(L⁶)_(q)-W⁶—R⁶;

wherein X is N or —C-(L)_(q)-W¹—R¹; or

wherein X is N or —C-(L²)_(q)-W²—R².

In an embodiment, the invention provides compounds having formula (FX7)to (FX12), where the variables are as defined above for formula (FX1):

In an embodiment, the invention provides a compound having formula(FX2), (FX5), (FX10) or (FX11).

In an embodiment, the invention provides compounds having formulas(FX13) to (FX17) where the variables are as defined above for formula(FX1):

wherein X is N or —C—W⁸—R⁸;

wherein X is N or —C—W⁷—R⁷;

wherein X is N or —C—W⁶—R⁶;

wherein X is N or —C—W¹—R¹;

wherein X is N or —C—W²—R².

L¹-L⁸ and W¹—W⁸ may be attaching and spacer groups, respectively, forproviding an appropriate linkage between R¹-R⁸ and the non-benzenoidaromatic ring core of compounds (FX1) to (FX17). In some embodiments,any or all of L¹-L⁸ or W¹-W⁸ may be present or absent. If a L¹-L⁸ orW¹-W⁸ is absent, there is a single bond between the substituents whichare present. For example, in an embodiment, the invention providescompounds useful as optical agents for phototherapeutic methods havingformula (FX1) wherein at least one of L¹-L⁸ is a single bond, andwherein at least one q is 0, thereby providing direct coupling of aW¹-W⁸, if present, or R¹-R⁸ to the backbone structure of the compound.In an embodiment, the invention provides compounds useful as opticalagents for phototherapeutic methods wherein at least one of W¹-W⁸ is asingle bond, providing direct coupling of at least one R¹-R⁸ to L¹-L⁸,if present, or to the backbone structure of the compound. In anembodiment, for example, the invention provides compounds useful asoptical agents for phototherapeutic methods having formula (FX1),wherein all q variables are 0, and wherein each of L¹-L⁸ is a singlebond.

In an embodiment, the invention provides compounds withelectron-donating and electron-withdrawing groups attached to adjacentpositions of the backbone. In an aspect of this embodiment, provided arecompounds of formula (FX1) to (FX17) wherein

(a) one of R¹ and R² is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R¹ and R² is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(b) one of R² and R³ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R² and R³ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(c) one of R³ and R⁴ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R³ and R⁴ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(d) one of R⁴ and R⁵ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R⁴ and R⁵ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(e) one of R⁵ and R⁶ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R⁵ and R⁶ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(f) one of R⁶ and R⁷ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R⁶ and R⁷ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl; or

(g) one of R⁷ and R⁸ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R⁷ and R⁸ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl.

In an embodiment, the invention provides compounds withelectron-donating and electron-withdrawing groups attached tonon-adjacent positions providing continuous conjugation between theelectron-donating and electron withdrawing groups. In an aspect of thisembodiment, the invention provides compounds of formula (FX1) to (FX17),wherein

(a) one of R¹ and R⁵ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R¹ and R⁵ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(b) one of R¹ and R⁷ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R and R⁷ is halo, trihalomethyl, —CN,—C₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —S₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₁-C₁₀acyl;

(c) one of R² and R⁴ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R² and R⁴ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(d) one of R² and R⁶ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R² and R⁶ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(e) one of R² and R⁸ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R² and R⁸ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl;

(f) one of R³ and R⁵ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R³ and R⁵ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl; or

(g) one of R⁴ and R⁷ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or—NR⁴⁸COR⁴⁹, and the other of R⁴ and R⁷ is halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, orC₁-C₁₀ acyl.

In an embodiment, the invention provides compounds of formula (FX1) to(FX17), wherein at least one of R¹ to R⁸ is C₁-C₆ alkyl, —OR⁴⁴, —SR⁴⁵,—NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹. In an embodiment, the invention providescompounds of formula (FX1) to (FX17), wherein at least one of R¹ to R⁸is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶,—SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₁-C₁₀ acyl. In an embodiment the inventionprovides compounds of formula (FX1) to (FX17), wherein at least one ofR¹ to R⁸ is TG¹ to TG⁸. In an embodiment the invention providescompounds of formula (FX1) to (FX17), wherein at least one of R¹ to R⁸is PS¹ to PS⁸. In an embodiment the invention provides compounds offormula (FX1) to (FX17), wherein at least one of R¹ to R⁸ is FL¹ to FL⁸.In an embodiment, the invention provides compounds of formula (FX1) to(FX17), wherein at least one of the substituent arms on rings A and Bcontains a polyethylene glycol (PEG) group. In embodiments, the PEGgroup provides increased water solubility over a compound that does notcontain a PEG group. In embodiments, the PEG group is abbreviated—(OCH₂CH₂)_(u)— where u is an integer from 1 to 25. The PEG group can beincorporated as a part of other substituent variables as providedherein.

In an embodiment, compounds of the invention include compounds havingthe numbering scheme described below,

wherein groups at one or more odd numbered positions 1, 3, 5, 7 and 9comprise hydrogen or electron withdrawing groups and groups at one ormore even numbered positions comprise hydrogen or halogen. Inembodiment, groups at one or more even numbered positions includehalogen. In an embodiment, groups at one or more even numbered groupsinclude fluorine.

In an embodiment, the invention provides compounds of formula (FX1) to(FX17), wherein the compound comprises at least one electron withdrawinggroup and at least one electron donating group. In an embodiment, theinvention provides compounds of formula (FX1) to (FX17), having R groupsubstituent pairings where one of the pair comprises anelectron-withdrawing group and the other of the pair comprises anelectron-donating group. In an aspect of this embodiment, the R groupsubstituent pairings are (R¹ and R²); (R² and R³); (R³ and R⁴); (R⁴ andR⁵); (R⁵ and R⁶); (R⁶ and R⁷); (R⁷ and R⁸); (R⁸ and R¹); (R¹ and R⁵),(R¹ and R⁷); (R² and R⁴); (R² and R⁶); (R² and R⁸); (R³ and R⁵); or (R⁴and R⁷), wherein one of the identified R groups in the substituentpairings is C₁-C₁₀ alkyl, —OR²⁸, —SR²⁹, —NR³⁰R³¹, or —NR³²COR³³ and theother of the identified R groups in the substituent pairings is halo,trihalomethyl, —CN, —CO₂R¹⁸, —CONR¹⁹R²⁰, —COR²¹, —NO₂, —SOR²², —SO₂R²⁴,or —SO₂NR²⁶R²⁷.

The invention includes, for example, compounds comprising one or morefluorescent moieties (or fluorophores). In an embodiment, the inventionprovides compounds of formulas (FX1) to (FX17) wherein at least one ofR¹ to R⁸ independently comprises a fluorescent moiety (abbreviated as FLherein). In an aspect of this embodiment, a fluorescent moiety is agroup corresponding to a naphthoquinone, an anthraquinone, anaphthacenedione, a pyrazine, an acridine, an acridone, aphenanthridine, a dibenzothiophene, a xanthene, a xanthone, a flavone, acoumarin, a phenoxazine, a phenothiazine, a phenoselenazine, a cyanine,an indocyanine, or an azo compound. The invention includes compoundshaving formulas (FX1) to (FX17) that do not include a FL group. Theinvention includes compounds having formulas (FX1) to (FX17) thatinclude a FL group and one or more of photosensitizing moiety ortargeting group.

The invention includes, for example, compounds having a photosensitizingmoiety (or photosensitizer) capable of producing one or more freeradicals, nitrenes, carbenes, and/or singlet oxygen. Methods for usingphotosensitizing moieties to produce free radicals, nitrenes, carbenes,and/or singlet oxygen are known in the art. In an embodiment, theinvention provides compounds of formulas (FX1) to (FX17) wherein atleast one of R¹ to R⁸ independently comprises a photosensitizing moiety(abbreviated as PS herein). In an aspect of this embodiment, aphotosensitizer moiety, if present, independently is a groupcorresponding to an azide, an azo, a diazo, an oxaza, a diaza, a thioxa,a phthalocyanine, a rhodamine, or a porphyrin group. The inventionincludes compounds having formulas (FX1) to (FX17) that do not include aPS group. The invention includes compounds having formulas (FX1) to(FX17) that include a PS group and one or more of fluorescent moiety ortargeting group.

The invention includes, for example, compounds having a targeting ligandor other molecular recognition component for delivering the opticalagent to a selected organ, tissue, or other cell material (targetinggroup). In an embodiment, the invention provides compounds of formulas(FX1) to (FX17), wherein at least one of R¹ to R⁸ independentlycomprises a targeting group (abbreviated as TG herein). In anembodiment, a targeting group, if present, is independently an aminoacid, a peptide, a protein, a nucleoside, a nucleotide, an enzyme, acarbohydrate, a glycomimetic, an oligomer, a lipid, a polymer, anantibody, an antibody fragment, a mono- or polysaccharide comprising 1to 50 carbohydrate units, a glycopeptide, a glycoprotein, apeptidomimetic, a drug, a drug mimic, a hormone, a receptor, a metalchelating agent, a radioactive or nonradioactive metal complex, a mono-or polynucleotide comprising 1 to 50 nucleic acid units, a polypeptidecomprising 2 to 30 amino acid units, or an echogenic agent. In anembodiment, a targeting group, if present, is independently an aminoacid, a peptide, a protein, a nucleoside, a nucleotide, an enzyme, anantibody, an antibody fragment, a saccharide, a glycopeptide, apeptidomimetic, a drug, a drug mimic, or a hormone. Incorporation of atargeting group or molecular recognition component in some compounds andmethods of the invention enables targeted delivery such that at least aportion of phototherapeutic agent administered to a subject accumulatesat a desired site, such as the site of an organ, tissue, tumor or otherlesion, prior to or during exposure to electromagnetic radiation. Theinvention includes compounds having formulas (FX1) to (FX17) that do nothave a TG group The invention includes compounds having formulas (FX1)to (FX17) that include a TG group and one or more of photosensitizingmoiety or fluorescent moiety. In an aspect of the present invention, acompound of the invention is targeted to a selected organ in thesubject. In an aspect of the present invention, a compound of theinvention is targeted to a selected tissue type in the subject. In anaspect of the present invention, the targeted tissue type is colon,renal, prostate, gastric, esophageal, uterine, endometrial, pancreatic,breast, cervical, brain, skin, gallbladder, lung, or ovary. In an aspectof the invention, the tissue type is renal.

In an embodiment, the invention provides compounds of formulas (FX1) to(FX17), wherein one of X_(a), X_(b), X_(c), X_(d) and X_(e) is N. In anembodiment, the invention provides compounds of formulas (FX1) to(FX17), wherein none of X_(a), X_(b), X_(c), X_(d) and X_(e) is N.

In an aspect of the present invention, the present invention is directedto a method of performing a medical imaging procedure on a patient. Themethod comprises administering to a subject an effective amount of acompound of the invention under conditions sufficient for contacting thecompound with a target cell; and exposing the administered compound toelectromagnetic radiation. In another aspect, the invention is directedto a method of performing a phototherapeutic procedure on a patient. Themethod comprises administering to a subject an effective amount of thecompound under conditions sufficient for contacting the compound withthe target cell; and exposing the administered compound toelectromagnetic radiation. In an aspect of the present invention, themedical imaging procedure or phototherapeutic procedure comprisesexposing the administered compound to electromagnetic radiation havingwavelengths selected over a range of 400 nanometers to 1300 nanometers.In an aspect of the present invention, the medical imaging procedure orphototherapeutic procedure comprises detecting electromagnetic radiationemitting from the compound in the subject. In an aspect of the medicalimaging procedure or phototherapeutic procedure of the presentinvention, exposing the compound administered to the subject toelectromagnetic radiation changes an optical property of the compound.In an aspect of the invention, the change in optical property of thecompound administered from exposure to electromagnetic radiation ismeasured or monitored. In an embodiment of this aspect of the invention,exposing the compound administered to the subject to electromagneticradiation increases the fluorescence intensity of the compound. In anembodiment of this aspect of the invention, exposing the compoundadministered to the subject to electromagnetic radiation changes theabsorption spectrum of the compound. In a further aspect, the presentinvention is directed to a method of performing a medical imagingprocedure on a patient, wherein the medical imaging procedure comprises:(a) administering to a subject an effective amount of the compound underconditions sufficient for contacting the compound with a target cell;(b) exposing the administered compound to electromagnetic radiation; and(c) detecting acoustic energy emitting from the compound in the subject.In embodiments, the compounds useful in the methods of the invention arecompounds of formulae (FX1) to (FX17). In embodiments, the compoundsuseful in the methods of the invention are compounds of formula (FX18):

wherein:

X_(a) is N or —C-(L⁶)_(q)-W⁶—R⁶;

X_(b) is N or —C-(L⁷)_(q)-W⁷—R⁷;

X_(c) is N or —C-(L⁸)_(q)-W⁸—R⁸;

X_(d) is N or —C-(L¹)_(q)-W¹—R¹;

X_(e) is N or —C-(L²)_(q)-W²—R²;

each of L¹ to L⁸ is independently C₁-C₁₀ alkylene, C₃-C₁₀ cycloalkylene,C₂-C₁₀ alkenylene, C₃-C₁₀ cycloalkenylene, C₂-C₁₀ alkynylene,ethenylene, ethynylene, phenylene, 1-aza-2,5-dioxocyclopentylene,—(CH₂CH₂O)_(m)—, —(CHOH)_(m)—, or 1,4-diazacyclohexylene;

each m is independently an integer selected from the range of 1 to 100;

each q is independently 0 or 1;

each of W¹ to W⁸ is independently a single bond, —(CH₂)_(n)—,—O(CH₂)_(n)—, —(CH₂)_(n)O—, —(HCCH)_(n)—, —O—, —S—, —SO—, —SO₂—, —SO₃ ⁻,—OSO₂—, —NR⁹—, —CO—, —COO—, —OCO—, —OCOO—, —CONR¹⁰—, —NR¹¹CO—,—OCONR¹²—, —NR¹³COO—, —NR¹⁴CONR¹⁵—, —NR¹⁶CSNR¹⁷—, —O(CH₂)_(n)—,—S(CH₂)_(n)—, —NR¹⁸(CH₂)_(n)—, —CO(CH₂)_(n)—, —COO(CH₂)_(n)—,—OCO(CH₂)_(n)—, —OCOO(CH₂)_(n)—, —CONR¹⁹(CH₂)_(n)—,—CONR²⁰(CH₂)_(n)(OCH₂CH₂)_(u)—, —NR²¹CO(CH₂)_(n)—, —OCONR²²(CH₂)_(n)—,—NR²³COO(CH₂)_(n)—, —NR²⁴CONR²⁵(CH₂)_(n)—, —NR²⁶CSNR²⁷(CH₂)_(n)—,—O(CH₂)_(n)NR²⁸CO(CH₂)_(n)—,—CO(CH₂)_(n)(CH₂OCH₂)_(n)(CH₂)_(n)NR²⁹(CH₂)_(n)NR³⁰CO—, —NR⁶⁹SR⁷⁰—, or—CO(CH₂)_(n)NR³¹CO—;

each n is independently an integer selected from the range of 1 to 10;

each of R⁹ to R³¹ and each of R⁶⁹ to R⁷⁰ is independently hydrogen,C₁-C₂₀ alkyl, or C₅-C₃₀ aryl;

each of R¹ to R⁸ is independently hydrogen, C₁-C₂₀ alkyl, C₅-C₃₀ aryl,C₁-C₂₀ acyl, C₂-C₂₀ alkenyl, C₂-C₂₀ alkynyl, C₅-C₂₀ alkylaryl, C₁-C₆alkoxycarbonyl, halo, halomethyl, dihalomethyl, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷, —SO₂R³⁸, —SO₂OR³⁹,—SO₂NR⁴⁰R⁴¹, —PO₃R⁴²R⁴³, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, —NR⁴⁸COR⁴⁹,—CH₂(CHOH)_(n)R⁵⁰, —(CH₂CH₂O)_(u)R⁵¹, —CH(R⁵²)CO₂H, —CH(R⁵³)NH₂, TG¹ toTG⁸, PS¹ to PS⁸, or FL¹ to FL⁸;

each u is independently an integer selected from the range of 1 to 25;

each of R³² to R⁵⁵ is independently hydrogen or C₁-C₁₀ alkyl;

each of TG¹ to TG⁸ is independently an amino acid, a peptide, a protein,a nucleoside, a nucleotide, an enzyme, a carbohydrate, a glycomimetic,an oligomer, a lipid, a polymer, an antibody, an antibody fragment, amono- or polysaccharide comprising 1 to 50 carbohydrate units, aglycopeptide, a glycoprotein, a peptidomimetic, a drug, a drug mimic, ahormone, a receptor, a metal chelating agent, a radioactive ornonradioactive metal complex, a mono- or polynucleotide comprising 1 to50 nucleic acid units, a polypeptide comprising 2 to 30 amino acidunits, or an echogenic agent;

each of PS¹ to PS⁸ is independently a photosensitizing moiety capable ofproducing one or more free radicals, nitrenes, carbenes, and/or singletoxygen, and wherein each of PS¹ to PS⁸ comprises at least one azide,azo, diazo, oxaza, diaza, dithia, thioxa, dioxa, phthalocyanine,rhodamine, or porphyrin group; and

each of FL¹ to FL⁸ is independently a fluorescent moiety selected fromnaphthoquinones, anthracenes, anthraquinones, phenanthrenes, tetracenes,naphthacenediones, pyridines, quinolines, isoquinolines, indoles,isoindoles, pyrroles, imidiazoles, pyrazoles, pyrazines, purines,benzimidazoles, benzofurans, dibenzofurans, carbazoles, acridines,acridones, phenanthridines, thiophenes, benzothiophenes,dibenzothiophenes, xanthenes, xanthones, flavones, coumarins,phenoxazines, phenothiazines, phenoselenazines, cyanines, indocyanines,and azo compounds;

wherein any adjacent R¹ to R⁸ may combine, optionally with one or two—CR⁵⁴R⁵⁵ groups, to form C₃-C₇ cycloalkyl, C₃-C₇ heterocycloalkyl, C₆aryl, or C₅-C₆ heteroaryl. In embodiments, where substituent groups andvarious aspects of formulae (FX1) to (FX17) are described, thesesubstituent groups and various aspects are intended to apply tocompounds of formula (FX18).

The present invention further includes compositions comprisingenantiomers, diastereomers and/or ionic forms (e.g., protonated anddeprotonated forms) of the compounds of formulae (FX1) to (FX17), andrelated methods of using compounds of formulae (FX1) to (FX17), forexample in a biomedical procedure. The present invention furtherincludes compositions comprising enantiomers, diastereomers and/or ionicforms (e.g., protonated and deprotonated forms) of the compounds offormula (FX18), and related methods of using compounds of formula(FX18), for example in a biomedical procedure.

In some embodiments, the present invention is directed to anon-benzenoid aromatic compound having at least one electron withdrawinggroup (EWG) and at least one electron donating group (EDG) bondeddirectly or indirectly to a carbon atom of the non-benzenoid ring core.In some embodiments, an electron withdrawing group and an electrondonating group are positioned on adjacent carbon atoms of thenon-benzenoid ring core. In some embodiments, an electron withdrawinggroup and an electron donating group are positioned on non-adjacentcarbon atoms of the non-benzenoid ring core. Multiple electronwithdrawing groups and/or electron donating groups bonded directly orindirectly to a carbon atom of the non-benzenoid ring core arecontemplated by this invention.

The present invention provides methods of making and using compounds,including compounds of formulas (FX1) to (FX18). Methods of this aspectof the present invention include in vivo, in vitro and ex vivo methodsfor biomedical and bioanalytical applications. Methods of the presentinvention include photodiagnostic and phototherapeutic methods, such asoptical imaging, anatomical visualization, endoscopic visualization,image guided surgery, and Type 1 and Type 2 phototherapy of tumors andother lesions. For some compounds for use in vivo, in vitro or ex vivofor imaging or visualizing, the tissue, organs and/or cells is a tumor,tumor site, or other lesion.

In an embodiment, the present invention provides pharmaceuticalcompositions of a therapeutically effective amount of one or morecompounds described herein, or their pharmaceutically acceptable salts.

Without wishing to be bound by any particular theory, there can bediscussion herein of beliefs or understandings of underlying principlesor mechanisms relating to the invention. It is recognized thatregardless of the ultimate correctness of any explanation or hypothesis,an embodiment of the invention can nonetheless be operative and useful.

Various features discussed herein in relation to one or more of theexemplary embodiments may be incorporated into any of the describedaspects of the present invention alone or in any combination. Certainexemplary aspects of the invention are set forth herein. It should beunderstood that these aspects are presented merely to provide the readerwith a brief summary of certain forms the invention might take and thatthese aspects are not intended to limit the scope of the invention.Indeed, the invention may encompass a variety of aspects that may not beexplicitly set forth herein as would be understood by one of ordinaryskill in the relevant art without undue experimentation.

DETAILED DESCRIPTION

In general, the terms and phrases used herein have their art-recognizedmeaning, which can be found by reference to standard texts, journalreferences and contexts known to those skilled in the art. The followingdefinitions are provided to clarify their specific use in the context ofthe invention.

The term “amino acid” comprises naturally occurring amino acids as wellas non-naturally occurring amino acids, including amino acid analogs andderivatives. One skilled in the art will recognize that reference hereinto an amino acid comprises, for example, naturally occurring proteogenicL-amino acids; D-amino acids; chemically modified amino acids such asamino acid analogs and derivatives; naturally occurring non-proteogenicamino acids, and chemically synthesized compounds having propertiesknown in the art to be characteristic of amino acids. Amino acidsinclude glycine, alanine, valine, leucine, isoleucine, methionine,proline, phenylalanine, tryptophan, asparagine, glutamine, glycine,serine, threonine, serine, rhreonine, asparagine, glutamine, tyrosine,cysteine, lysine, arginine, histidine, aspartic acid and glutamic acid.As used herein, reference to “a side chain residue of a natural α-aminoacid” specifically includes the side chains of the above-referencedamino acids.

The term “nucleic acid” as used herein generally refers to a molecule orstrand of DNA, RNA, or derivatives or analogs thereof including one ormore nucleobases. Nucleobases comprise purine or pyrimidine basestypically found in DNA or RNA (e.g., adenine, guanine, thymine,cytosine, and/or uracil). The term “nucleic acid” also comprisesoligonucleotides and polynucleotides. Nucleic acids may besingle-stranded molecules, or they may be double-, triple- orquadruple-stranded molecules that may comprise one or more complementarystrands of a particular molecule. “Nucleic acid” includes artificialnucleic acids including peptide nucleic acids, morpholino nucleic acids,glycol nucleic acids and threose nucleic acids. Artificial nucleic acidsmay be capable of nucleic acid hybridization.

As used herein, “sequence” means the linear order in which monomersoccur in a polymer, the order of amino acids in a polypeptide or theorder of nucleotides in a polynucleotide for example.

The terms “peptide” and “polypeptide” are used synonymously in thepresent description, and refer to a class of compounds composed of aminoacid residues chemically bonded together by amide bonds (or peptidebonds), regardless of length, functionality, environment, or associatedmolecule(s) Peptides and polypeptides are polymeric compounds comprisingat least two amino acid residues or modified amino acid residues.Modifications can be naturally occurring or non-naturally occurring,such as modifications generated by chemical synthesis. Modifications toamino acids in peptides include, but are not limited to,phosphorylation, glycosylation, lipidation, prenylation, sulfonation,hydroxylation, acetylation, methionine oxidation, alkylation, acylation,carbamylation, iodination and the addition of cofactors. Peptidesinclude proteins and further include compositions generated bydegradation of proteins, for example by proteolyic digestion. Peptidesand polypeptides can be generated by substantially complete digestion orby partial digestion of proteins. Polypeptides comprising 2 to 100 aminoacid units, optionally for some embodiments 2 to 50 amino acid unitsand, optionally for some embodiments 2 to 20 amino acid units can beused as polypeptide targeting ligands in the invention, for example,where the polypeptide preferentially binds to proteins, peptides orother biomolecules expressed, or otherwise generated by, a targettissue, such as a tumor, precancerous tissue, site of inflammation orother lesion. Typically, the polypeptide is at least four amino acidresidues in length and can range up to a full-length protein.

“Target tissue” refers to tissue of a subject to which an optical agentis administered or otherwise contacted, for example during a biomedicalprocedure such as an optical imaging, phototherapy, monitoring orvisualization procedure. Target tissues can be contacted with an opticalagent of the invention under in vivo conditions or ex vivo conditions.Target tissues in some embodiments include cancerous tissue, cancercells, precancerous tissue, a tumor, a lesion, a site of inflammation,or vasculature tissue. In some embodiments, a target tissue includes amelanoma cell, a breast lesion, a prostate lesion, a lung cancer cell, acolorectal cancer cell, an atherosclerotic plaque, a brain lesion, ablood vessel lesion, a lung lesion, a heart lesion, a throat lesion, anear lesion, a rectal lesion, a bladder lesion, a stomach lesion, anintestinal lesion, an esophagus lesion, a liver lesion, a pancreaticlesion, and a solid tumor. Target tissue in some embodiments refers to aselected organ of the subject or component thereof, such as lung, heart,brain, stomach, liver, kidneys, gallbladder, pancreas, intestines,rectum, skin, colon, prostate, ovaries, breast, bladder, blood vessel,throat, ear, or esophagus.

“Protein” refers to a class of compounds comprising one or morepolypeptide chains and/or modified polypeptide chains. Proteins can bemodified by naturally occurring processes such as post-translationalmodifications or co-translational modifications. Exemplarypost-translational modifications or co-translational modificationsinclude, but are not limited to, phosphorylation, glycosylation,lipidation, prenylation, sulfonation, hydroxylation, acetylation,methionine oxidation, the addition of cofactors, proteolysis, andassembly of proteins into macromolecular complexes. Modification ofproteins can also include non-naturally occurring derivatives, analoguesand functional mimetics generated by chemical synthesis. Exemplaryderivatives include chemical modifications such as alkylation,acylation, carbamylation, iodination or any modification thatderivatizes the protein.

As used herein, “polynucleotide” and “oligonucleotide” are usedinterchangeably and refer to a class of compounds composed of nucleicacid residues chemically bonded together. The invention provides opticalagents having an oligonucleotide or polynucleotide targeting ligandwhich comprises a plurality of nucleic acid residues, such as DNA or RNAresidues, and/or modified nucleic acid residues that preferentiallybinds to proteins, peptides or other biomolecules expressed, orotherwise generated by, a target tissue, such as a tumor, precanceroustissue, site of inflammation or other lesion. Modifications to nucleicacid residues can be naturally occurring or non-naturally occurring,such as modifications generated by chemical synthesis. Oligo- orpoly-nucleotide targeting ligands include, for example, oligo- orpoly-nucleotides comprising 1 to 100 nucleic acid units, optionally forsome embodiments 1 to 50 nucleic acid units and, optionally for someembodiments 1 to 20 nucleic acid units. Polypeptide and oligonucleotideinclude a polymer of at least two nucleotides joined together byphosphodiester bonds and may consist of either ribonucleotides ordeoxyribonucleotides.

“Peptidomimetic” refers to a small molecule having activity, includingbiological activity that resembles that of a polypeptide. Morphine, forexample, is a peptidomimetic of endorphin peptide. In some embodiments,a peptidomimetic is a small protein-like polymer designed to mimic thefunctionality of a peptide. Peptidomimetics useful as targeting ligandsfor some compounds of the invention in the present invention includepeptoids and β-peptides.

The term “carbocyclic” refers to ring structures containing only carbonatoms in the ring. Carbon atoms of carbocyclic rings can be bonded to awide range of other atoms and functional groups.

The term “alicyclic” refers to a ring that is not an aromatic ring.Alicyclic rings include both carbocyclic and heterocyclic rings.

The term “heterocyclic” refers to ring structures containing at leastone other kind of atom, in addition to carbon, in the ring. Examples ofsuch atoms include nitrogen, oxygen and sulfur. Examples of heterocyclicrings include, but are not limited to, pyrrolidinyl, piperidyl,imidazolidinyl, tetrahydrofuryl, tetrahydrothienyl, furyl, thienyl,pyridyl, quinolyl, isoquinolyl, pyridazinyl, pyrazinyl, indolyl,imidazolyl, oxazolyl, thiazolyl, pyrazolyl, pyridinyl, benzoxadiazolyl,benzothiadiazolyl, triazolyl and tetrazolyl groups.

As used herein, the term “alkoxyalkyl” refers to a substituent of theformula alkyl-O-alkyl.

As used herein, the term “polyhydroxyalkyl” refers to a substituenthaving from 2 to 12 carbon atoms and from 2 to 5 hydroxyl groups, suchas the 2,3-dihydroxypropyl, 2,3,4-trihydroxybutyl or2,3,4,5-tetrahydroxypentyl residue.

As used herein, the term “polyalkoxyalkyl” refers to a substituent ofthe formula alkyl-(alkoxy)_(n)-alkoxy wherein n is an integer from 1 to10, preferably 1 to 4, and more preferably for some embodiments 1 to 3.

As used herein, a “non-benzenoid aromatic” compound or derivative is anaromatic compound having two or more fused non-benzene rings as a corestructure. Examples of non-benzenoid aromatic compounds include azuleneand azaazulene. Non-benzenoid aromatic compounds are also referred to asnon-benzenoid compounds herein.

“Optical agent” generally refers to compounds, compositions,preparations, and/or formulations that absorb, emit, or scatterelectromagnetic radiation of wavelength, generally in the range of400-900 nanometers, within a biologically relevant environment orcondition. Optical agents optionally have molecular recognition ortargeting functions enabling localized delivery to a target tissue. Insome embodiments, optical agents of the present invention, when excitedby electromagnetic radiation, undergo emission via fluorescence orphosphorescence pathways. These pathways are useful for diagnosticimaging, visualization, or organ function monitoring. Compoundsbelonging to this class are commonly referred to as “optical imagingagents” or “optical contrast agents.” In some other embodiments, opticalagents of the present invention absorb electromagnetic radiation andundergo photochemical reactions such as photofragmentation of one ormore photolabile bonds to generate reactive intermediates such asnitrenes, carbine, free radicals, or ions. This process is useful forphototherapy of tumors or other lesions. Compounds belonging to thisclass are commonly referred to as “photosensitizers.”

Optical agents of the present invention include, but are not limited to,contrast agents, imaging agents, dyes, detectable agents,photosensitizer agents, photoactivators, and photoreactive agents; andconjugates, complexes, and derivatives thereof. Optical agents of thepresent invention include non-benzenoid derivatives having anon-benzenoid ring core structure and derivatives thereof. Some opticalagents of the present invention provide detectable agents that can beadministered to a subject and subsequently detected using a variety ofoptical techniques, including optical imaging, visualization, and otherforms of optical detection.

Optical agents of the present invention can contain fluorophores. Theterm “fluorophore” generally refers to a component or moiety of amolecule or group which causes a molecule or group to be fluorescent.Fluorophores can be functional groups in a molecule which absorbelectromagnetic radiation of first specific wavelengths and re-emitenergy at second specific wavelengths. The amount and wavelengths of theemitted electromagnetic radiation depend on both the fluorophore and thechemical environment of the fluorophore. The term “fluorophore” isabbreviated throughout the present description as “FL”. In aspects ofthe invention, fluorophores emit energy in the visible (e.g. 350 nm to750 nm) and NIR regions (e.g., 750-1300 nm) of the electromagneticspectrum.

As used herein, a “chromophore” is a compound or functional group of acompound that absorbs electromagnetic radiation, preferably for someapplications electromagnetic radiation having wavelengths in the UV(e.g. 200 nm to 350 nm) or visible (e.g. 350 nm to 750 nm) of theelectromagnetic spectrum.

As used herein, an “electron withdrawing group” (EWG) refers to anychemical group that draws electrons from a center, such as thenon-benzenoid core of the present invention. In an embodiment, electronwithdrawing group(s) as substituent groups for the compositions offormulae (FX1) to (FX18) are independently selected from the groupconsisting of: cyano (—CN), carbonyl (—CO), carboxylate (—CO₂R⁵⁰),carbamate (—CONR⁵¹R⁵²), halo (—F, —Cl, —Br, —I, —At), acyl (—COR⁵³),nitro (—NO₂), sulfinyl (—SOR⁵⁴), —OSR⁵⁵, sulfonyl (—SO₂R⁵⁶, —SO₂OR⁵⁷,and —SO₂NR⁵⁸R⁵⁹), and —PO₃R⁶⁰R⁶¹, wherein R⁵⁰-R⁶¹ are in some instancesindependently selected to enhance biological and/or physiochemicalproperties of the non-benzenoid derivatives of the present invention. Insome instances, R⁵⁰-R⁶¹ are independently any one of a hydrogen atom, ananionic functional group (e.g., carboxylate, sulfonate, sulfate,phosphonate or phosphate) or a hydrophilic functional group (e.g.,hydroxyl, carboxyl, sulfonyl, sulfonato or phosphonato). In otherinstances, R⁵⁰-R⁶¹ are independently hydrogen, C₁₋₁₀ alkyl, aryl,heteroaryl, —(CH₂)_(a)OH, —(CH₂)_(a)CO₂H, —(CH₂)_(a)SO₃H, —(CH₂)_(a)SO₃⁻, —(CH₂)_(a)OSO₃H, —(CH₂)_(a)OSO₃ ⁻, —(CH₂)_(a)NHSO₃H, —(CH₂)_(a)NHSO₃⁻, —(CH₂)_(a)PO₃H₂, —(CH₂)_(a)PO₃H—, —(CH₂)_(a)PO₃ ²⁻, —(CH₂)_(a)OPO₃H₂,—(CH₂)_(a)OPO₃H— or —(CH₂)_(a)OPO₃ where a is an integer from 1 to 10.In one example of this embodiment, the EWG(s) are independently —CN,—CO₂R⁵⁰, —CONR⁵¹R⁵², —COR⁵³, —NO₂, or —SO₂R⁵⁶. In an embodiment, an EWGis located at the terminus of a substituent arm of a non-benzenoidaromatic compound of the present invention.

As used herein, an “electron donating group” (EDG) refers to anychemical group that releases electrons to a center, such as thenon-benzenoid core of the present invention. In an embodiment, electrondonating group(s) as substituent groups for the compositions of formulae(FX1) to (FX18) are independently selected from the group consisting of:C₁-C₁₀ alkyl, C₅-C₁₀ aryl, —(CH₂)_(n)OH—OR⁶², —SR⁶³, —NR⁶⁴R⁶⁵,—N(R⁶⁶)COR⁶⁷, and —P(R⁶⁸) wherein R⁶²-R⁶⁸ are in some instancesindependently selected to enhance biological and/or physiochemicalproperties of the non-benzenoid core of the present invention. n isselected from the range of 1 to 10. In some instances, R⁶²-R⁶⁸ areindependently any one of a hydrogen atom, an anionic functional group(e.g., carboxylate, sulfonate, sulfate, phosphonate and phosphate) or ahydrophilic functional group (e.g., hydroxyl, carboxyl, sulfonyl,sulfonato and phosphonato). In other instances, R⁶²-R⁶⁸ areindependently hydrogen, C₁₋₁₀ alkyl, aryl, heteroaryl, —(CH₂)_(a)OH,—(CH₂)_(a)CO₂H, —(CH₂)_(a)SO₃H, —(CH₂)_(a)SO₃ ⁻, —(CH₂)_(a)OSO₃H,—(CH₂)_(a)OSO₃ ⁻, —(CH₂)_(a)NHSO₃H, —(CH₂)_(a)NHSO₃ ⁻, —(CH₂)_(a)PO₃H₂,—(CH₂)_(a)PO₃H—, —(CH₂)_(a)PO₃ ²⁻, —(CH₂)_(a)OPO₃H₂, —(CH₂)_(a)OPO₃H— or—(CH₂)_(a)OPO₃ where a is an integer from 1 to 10. In one example ofthis embodiment, the EDG(s) are independently —OR⁶², —SR⁶³, —NR⁶⁴R⁶⁵, or—N(R⁶⁶)COR⁶⁷. In an embodiment, an EDG is located at the terminus of asubstituent arm of a non-benzenoid aromatic compound of the presentinvention.

As used throughout the present description, the expression “a groupcorresponding to” an indicated species expressly includes a radical,including monovalent, divalent and polyvalent radicals. for example, anaromatic or heterocyclic aromatic radical, of the groups listed providedin a covalently bonded configuration, optionally with one or moresubstituents, including but not limited to electron donating groups,electron withdrawing groups and/or targeting ligands.

In an embodiment, an effective amount of a compound or composition ofthe invention is a therapeutically effective amount. As used herein, thephrase “therapeutically effective” qualifies the amount of compoundadministered in the therapy. This amount achieves the goal ofameliorating, suppressing, eradicating, preventing, reducing the riskof, or delaying the onset of the targeted condition.

In an embodiment, an effective amount of a compound or composition ofthe invention is a diagnostically effective amount. As used herein, thephrase “diagnostically effective” qualifies the amount of compoundadministered in diagnosis. The amount achieves the goal of beingdetectable while avoiding adverse side effects found with higher doses.In an embodiment, an active ingredient or other component is included ina therapeutically acceptable amount. In an embodiment, an activeingredient or other component is included in a diagnostically acceptableamount.

“Photosensitizing moiety”, “photosensitizers” and “phototherapeuticagents” are used interchangeably and refer to a class of optical agentsthat absorb electromagnetic radiation and undergo photochemicalreactions, such as photofragmentation of one or more photolabile bonds,to generate reactive intermediates for achieving a desired therapeuticresult. Phototherapeutic agents include compounds that absorb visibleand/or near infrared radiation and generate one or more nitrenes,carbene, free radicals, singlet oxygen and/or ions. Phototherapeuticagents are useful for a wide range of phototherapy applications, forexample in the treatment of tumors or other lesions. Photosensitizersinclude Type 1 and Type 2 phototherapeutic agents.

“Optical condition” refers to one or more of the following: thefluorescence quantum yield, fluorescence intensity, fluorescenceexcitation wavelength, wavelength distribution or spectrum, emissionwavelength, wavelength distribution or spectrum, Stokes shift, color,reflectance, phosphorescence, chemiluminescence, scattering, and/orother observable and/or measurable spectral property or phenomenon.

“Phototherapy procedure” refers to a therapeutic procedure involvingadministration of a phototherapeutic agent compound of the inventionwhich comprises a photosensitizing moiety to a patient followed bysubsequent excitation of the phototherapeutic agent by exposure toapplied electromagnetic radiation, such as electromagnetic radiationhaving wavelengths in the visible and/or near IR region of theelectromagnetic spectrum, such as wavelengths in the range of 400-1300nanometers. Phototherapy includes, but is not limited to, photodynamictherapy. Phototherapeutic agents of the invention can also comprise oneor more of targeting groups, fluorescent moieties, electron withdrawinggroups, electron donating groups, and other groups.

As used herein, the term “pharmaceutically acceptable salt” can refer toacid addition salts or base addition salts of the compounds in thepresent disclosure. A pharmaceutically acceptable salt is any salt whichretains the activity of the parent compound and does not impart anydeleterious or undesirable effect on a subject to whom it isadministered and in the context in which it is administered.Pharmaceutically acceptable salts include metal complexes and salts ofboth inorganic and organic acids. Pharmaceutically acceptable saltsinclude metal salts such as aluminum, calcium, iron, magnesium,manganese and complex salts. Pharmaceutically acceptable salts include,but are not limited to, acid salts such as acetic, aspartic,alkylsulfonic, arylsulfonic, axetil, benzenesulfonic, benzoic,bicarbonic, bisulfuric, bitartaric, butyric, calcium edetate, camsylic,carbonic, chlorobenzoic, -32-cilexetil, citric, edetic, edisylic,estolic, esyl, esylic, formic, fumaric, gluceptic, gluconic, glutamic,glycolic, glycolylarsanilic, hexamic, hexylresorcjnoic, hydrabamic,hydrobromic, hydrochloric, hydroiodic, hydroxynaphthoic, isethionic,lactic, lactobionic, maleic, malic, malonic, mandelic, methanesulfonic,methylnitric, methylsulfuric, mucic, muconic, napsylic, nitric, oxalic,p-nitromethanesulfonic, pamoic, pantothenic, phosphoric, monohydrogenphosphoric, dihydrogen phosphoric, phthalic, polygalactouronic,propionic, salicylic, stearic, succinic, sulfamic, sulfanlic, sulfonic,sulfuric, tannic, tartaric, teoclic, toluenesulfonic, and the like.Pharmaceutically acceptable salts may be derived from amino acids,including but not limited to cysteine. Other pharmaceutically acceptablesalts may be found, for example, in Stahl et al., Handbook ofPharmaceutical Salts: Properties, Selection, and Use, Wiley-VCH; VerlagHelvetica Chimica Acta, Zurich, 2002. (ISBN 3-906390-26-8).

When used herein, the term “diagnosis”, “diagnostic” and other root wordderivatives are as understood in the art and are further intended toinclude a general monitoring, characterizing and/or identifying a stateof health, physical state, or disease. The term is meant to encompassthe concept of prognosis. For example, the diagnosis of cancer caninclude an initial determination and/or one or more subsequentassessments regardless of the outcome of a previous finding. The termdoes not necessarily imply a defined level of certainty regarding theprediction of a particular status or outcome.

As used herein, “administering” means that a compound or formulationthereof of the present invention, such as a non-benzenoid aromaticcompound, is provided to a patient or subject, for example in atherapeutically effective amount. “Co-administration” refers toadministering two or more compounds at some time during a biomedicalprocedure. Co-administration refers to administration of two or morecompounds at the same time, or before or after each other during thesame biomedical procedure. Co-administration includes phototherapyprocedures wherein a non-benzenoid aromatic compound is administeredprior to excitation of a phototherapeutic agent, during excitation ofthe phototherapeutic agent and/or after excitation of a phototherapeuticagent. Co-administration can include multiple administrations during thesame biomedical procedure.

Alkyl groups include straight-chain, branched and cyclic (e.g.,cycloalkyl) alkyl groups. Alkyl groups include those having from 1 to 30carbon atoms. Alkyl groups include small alkyl groups having 1 to 3carbon atoms. Alkyl groups include medium length alkyl groups havingfrom 4-10 carbon atoms. Alkyl groups include long alkyl groups havingmore than 10 carbon atoms, particularly those having 10-30 carbon atoms.Cyclic alkyl groups include those having one or more rings. Cyclic alkylgroups include those having a 3-, 4-, 5-, 6-, 7-, 8-, 9- or 10-membercarbon ring and particularly those having a 3-, 4-, 5-, 6-, or 7-memberring. The carbon rings in cyclic alkyl groups can also carry alkylgroups. Cyclic alkyl groups can include bicyclic and tricyclic alkylgroups. Alkyl groups are optionally substituted. Substituted alkylgroups include among others those which are substituted with arylgroups, which in turn can be optionally substituted. Specific alkylgroups include methyl, ethyl, n-propyl, iso-propyl, cyclopropyl,n-butyl, s-butyl, t-butyl, cyclobutyl, n-pentyl, branched-pentyl,cyclopentyl, n-hexyl, branched hexyl, and cyclohexyl groups, all ofwhich are optionally substituted. Substituted alkyl groups include fullyhalogenated or semihalogenated alkyl groups, such as alkyl groups havingone or more hydrogens replaced with one or more fluorine atoms, chlorineatoms, bromine atoms and/or iodine atoms. Substituted alkyl groupsinclude fully fluorinated or semifluorinated alkyl groups, such as alkylgroups having one or more hydrogens replaced with one or more fluorineatoms. An alkoxy group is an alkyl group linked to oxygen and can berepresented by the formula R—O. Examples of alkoxy groups include, butare not limited to, methoxy, ethoxy, propoxy, butoxy and heptoxy. Alkoxygroups include substituted alkoxy groups wherein the alkyl portion ofthe groups is substituted as provided herein in connection with thedescription of alkyl groups. As used herein MeO— refers to CH₃O—.

Alkenyl groups include straight-chain, branched and cyclic (e.g.,cycloalkenyl) alkenyl groups. Alkenyl groups include those having 1, 2or more double bonds and those in which two or more of the double bondsare conjugated double bonds. Alkenyl groups include those having from 2to 20 carbon atoms. Alkenyl groups include small alkenyl groups having 2to 3 carbon atoms. Alkenyl groups include medium length alkenyl groupshaving from 4-10 carbon atoms. Alkenyl groups include long alkenylgroups having more than 10 carbon atoms, particularly those having 10-20carbon atoms. Cyclic alkenyl groups include those having one or morerings. Cyclic alkenyl groups include those in which a double bond is inthe ring or in an alkenyl group attached to a ring. Cyclic alkenylgroups include those having a 3-, 4-, 5-, 6-, 7-, 8-, 9- or 10-membercarbon ring and particularly those having a 3-, 4-, 5-, 6- or 7-memberring. The carbon rings in cyclic alkenyl groups can also carry alkylgroups. Cyclic alkenyl groups can include bicyclic and tricyclic alkylgroups. Alkenyl groups are optionally substituted. Substituted alkenylgroups include among others those which are substituted with alkyl oraryl groups, which groups in turn can be optionally substituted.Specific alkenyl groups include ethenyl, prop-1-enyl, prop-2-enyl,cycloprop-1-enyl, but-1-enyl, but-2-enyl, cyclobut-1-enyl,cyclobut-2-enyl, pent-1-enyl, pent-2-enyl, branched pentenyl,cyclopent-1-enyl, hex-1-enyl, branched hexenyl, cyclohexenyl, all ofwhich are optionally substituted. Substituted alkenyl groups includefully halogenated or semihalogenated alkenyl groups, such as alkenylgroups having one or more hydrogens replaced with one or more fluorineatoms, chlorine atoms, bromine atoms and/or iodine atoms. Substitutedalkenyl groups include fully fluorinated or semifluorinated alkenylgroups, such as alkenyl groups having one or more hydrogens replacedwith one or more fluorine atoms.

Aryl groups include groups having one or more 5- or 6- or 7-memberaromatic or heteroaromatic rings. Heteroaryl groups are aryl groupshaving one or more heteroatoms (N, O or S) in the ring. Aryl groups cancontain one or more fused aromatic rings or a combination of one or morearomatic or heteroaromatic rings and one or more nonaromatic rings thatmay be fused or linked via covalent bonds. Heterocyclic aromatic ringscan include one or more N, O, or S atoms in the ring. Heterocyclicaromatic rings can include those with one, two or three N, those withone or two O, and those with one or two S, or combinations of one or twoor three N, O or S. Aryl groups are optionally substituted. Substitutedaryl groups include among others those which are substituted with alkylor alkenyl groups, which groups in turn can be optionally substituted.Specific aryl groups include phenyl groups, biphenyl groups, pyridinylgroups, and naphthyl groups, all of which are optionally substituted.Substituted aryl groups include fully halogenated or semihalogenatedaryl groups, such as aryl groups having one or more hydrogens replacedwith one or more fluorine atoms, chlorine atoms, bromine atoms and/oriodine atoms. Substituted aryl groups include fully fluorinated orsemifluorinated aryl groups, such as aryl groups having one or morehydrogens replaced with one or more fluorine atoms. Aryl groups include,but are not limited to, aromatic group-containing or heterocylicaromatic group-containing groups corresponding to any one of thefollowing: benzene, naphthalene, naphthoquinone, diphenylmethane,fluorene, anthracene, anthraquinone, phenanthrene, tetracene,tetracenedione, pyridine, quinoline, isoquinoline, indoles, isoindole,pyrrole, imidazole, oxazole, thiazole, pyrazole, pyrazine, pyrimidine,purine, benzimidazole, furans, benzofuran, dibenzofuran, carbazole,acridine, acridone, phenanthridine, thiophene, benzothiophene,dibenzothiophene, xanthene, xanthone, flavone, coumarin, azulene oranthracycline. As used herein, a group corresponding to the groupslisted above expressly includes an aromatic or heterocyclic aromaticradical, including monovalent, divalent and polyvalent radicals, of thearomatic and heterocyclic aromatic groups listed herein are provided ina covalently bonded configuration in the compounds of the invention atany suitable point of attachment. In embodiments, aryl groups containbetween 5 and 30 carbon atoms. In embodiments, aryl groups contain onearomatic or heteroaromatic six-membered ring and one or more additionalfive- or six-membered aromatic or heteroaromatic ring. In embodiments,aryl groups contain between five and eighteen carbon atoms in the rings.Aryl groups optionally have one or more aromatic rings or heterocyclicaromatic rings having one or more electron donating groups, electronwithdrawing groups and/or targeting ligands provided as substituents.Aryl groups include the following compounds and compounds including thefollowing compounds:

Arylalkyl groups or alkylaryl groups are alkyl groups substituted withone or more aryl groups wherein the alkyl groups optionally carryadditional substituents and the aryl groups are optionally substituted.Specific alkylaryl groups are phenyl-substituted alkyl groups, e.g.,phenylmethyl groups. Alkylaryl groups are alternatively described asaryl groups substituted with one or more alkyl groups wherein the alkylgroups optionally carry additional substituents and the aryl groups areoptionally substituted. Specific alkylaryl groups are alkyl-substitutedphenyl groups such as methylphenyl. Substituted arylalkyl groups includefully halogenated or semihalogenated arylalkyl groups, such as arylalkylgroups having one or more alkyl and/or aryl having one or more hydrogensreplaced with one or more fluorine atoms, chlorine atoms, bromine atomsand/or iodine atoms.

Optional substitution of any alkyl, alkenyl, and aryl groups includessubstitution with one or more of the following substituents: halogens,—CN, —COOR, —OR, —COR, —OCOOR, —CON(R)₂, —OCON(R)₂, —N(R)₂, —NO₂, —SR,—SO₂R, —SO₂N(R)₂ or —SOR groups. Optional substitution of alkyl groupsincludes substitution with one or more alkenyl groups, aryl groups orboth, wherein the alkenyl groups or aryl groups are optionallysubstituted. Optional substitution of alkenyl groups includessubstitution with one or more alkyl groups, aryl groups, or both,wherein the alkyl groups or aryl groups are optionally substituted.Optional substitution of aryl groups includes substitution of the arylring with one or more alkyl groups, alkenyl groups, or both, wherein thealkyl groups or alkenyl groups are optionally substituted.

Optional substituents for alkyl, alkenyl and aryl groups include amongothers:

—COOR where R is a hydrogen or an alkyl group or an aryl group and morespecifically where R is methyl, ethyl, propyl, butyl, or phenyl groupsall of which are optionally substituted;

—COR where R is a hydrogen, or an alkyl group or an aryl group and morespecifically where R is methyl, ethyl, propyl, butyl, or phenyl groupsall of which groups are optionally substituted; —CON(R)₂ where each R,independently of each other R, is a hydrogen or an alkyl group or anaryl group and more specifically where R is methyl, ethyl, propyl,butyl, or phenyl groups all of which groups are optionally substituted;R and R can form a ring which may contain one or more double bonds;—OCON(R)₂ where each R, independently of each other R, is a hydrogen oran alkyl group or an aryl group and more specifically where R is methyl,ethyl, propyl, butyl, or phenyl groups all of which groups areoptionally substituted; R and R can form a ring which may contain one ormore double bonds;—N(R)₂ where each R, independently of each other R, is a hydrogen, or analkyl group, acyl group or an aryl group and more specifically where Ris methyl, ethyl, propyl, butyl, or phenyl or acetyl groups all of whichare optionally substituted; or R and R can form a ring which may containone or more double bonds.—SR, —SO₂R, or —SOR where R is an alkyl group or an aryl groups and morespecifically where R is methyl, ethyl, propyl, butyl, phenyl groups allof which are optionally substituted; for —SR, R can be hydrogen;—OCOOR where R is an alkyl group or an aryl group;—SO₂N(R)₂ where R is a hydrogen, an alkyl group, or an aryl group and Rand R can form a ring;—OR where R═H, alkyl, aryl, or acyl; for example, R can be an acylyielding —OCOR* where R* is a hydrogen or an alkyl group or an arylgroup and more specifically where R* is methyl, ethyl, propyl, butyl, orphenyl groups all of which groups are optionally substituted.

Specific substituted alkyl groups include haloalkyl groups, particularlytrihalomethyl groups and specifically trifluoromethyl groups. Specificsubstituted aryl groups include mono-, di-, tri, tetra- andpentahalo-substituted phenyl groups; mono-, di-, tri-, tetra-, penta-,hexa-, and hepta-halo-substituted naphthalene groups; 3- or4-halo-substituted phenyl groups, 3- or 4-alkyl-substituted phenylgroups, 3- or 4-alkoxy-substituted phenyl groups, 3- or4-RCO-substituted phenyl, 5- or 6-halo-substituted naphthalene groups.More specifically, substituted aryl groups include acetylphenyl groups,particularly 4-acetylphenyl groups; fluorophenyl groups, particularly3-fluorophenyl and 4-fluorophenyl groups; chlorophenyl groups,particularly 3-chlorophenyl and 4-chlorophenyl groups; methylphenylgroups, particularly 4-methylphenyl groups, and methoxyphenyl groups,particularly 4-methoxyphenyl groups. As used herein, the term“polyhydroxylalkyl” refers to an alkyl group having more than onehydroxy substitution.

As used herein, the term “alkylene” refers to a divalent radical derivedfrom an alkyl group as defined herein. Alkylene groups in someembodiments function as bridging and/or spacer groups in the presentcompositions. Compounds of the invention include substituted andunsubstituted C1-C20 alkylene, C1-C10 alkylene and C1-C5 alkylenegroups.

As used herein, the term “arylene” refers to a divalent radical derivedfrom an aryl group as defined herein. Arylene groups in some embodimentsfunction as attaching and/or spacer groups in the present compositions.Arylene groups in other embodiments function as dye and/or imaginggroups in the present compositions. Compounds of the invention includesubstituted and unsubstituted C₁-C₂₀ arylene, C₁-C₁₀ arylene and C₁-C₅arylene groups.

As used herein, the term “cycloalkylene” refers to a divalent radicalderived from a cycloalkyl group as defined herein. Cycloalkylene groupsin some embodiments function as bridging and/or spacer groups in thepresent compositions. Compounds of the invention include substituted andunsubstituted C1-C20 cycloalkylene, C1-C10 cycloalkylene and C1-C5cycloalkylene groups.

As used herein, the term “alkenylene” refers to a divalent radicalderived from an alkenyl group as defined herein. Alkenylene groups insome embodiments function as bridging and/or spacer groups in thepresent compositions. Compounds of the invention include substituted andunsubstituted C1-C20 alkenylene, C1-C10 alkenylene and C1-C5 alkenylenegroups.

As used herein, the term “cycloalkenylene” refers to a divalent radicalderived from a cycloalkenyl group as defined herein. Cycloalkenylenegroups in some embodiments function as bridging and/or spacer groups inthe present compositions. Compounds of the invention include substitutedand unsubstituted C1-C20 cycloalkenylene, C1-C10 cycloalkenylene andC1-C5 cycloalkenylene groups.

As used herein, the term “alkynylene” refers to a divalent radicalderived from an alkynyl group as defined herein. Alkynylene groups insome embodiments function as bridging and/or spacer groups in thepresent compositions. Compounds of the invention include substituted andunsubstituted C1-C20 alkynylene, C1-C10 alkynylene and C1-C5 alkynylenegroups.

As used herein, the term “halo” or “halogen” refers to a halogen groupsuch as a fluoro (—F), chloro (—Cl), bromo (—Br) or iodo (—I).

As is customary and well known in the art, hydrogen atoms in theformulas shown herein, including formulae (FX1)-(FX18) are not alwaysexplicitly shown. The structures provided herein, for example in thecontext of the description of formulas (FX1)-(FX18), are intended toconvey to one of reasonable skill in the art the chemical composition ofcompounds of the methods and compositions of the invention, and as willbe understood by one of skill in the art, the structures provided do notindicate the specific bond angles between atoms of these compounds.

As used herein, the term “echogenic agent” is used as conventional inthe art and is generally a compound used in connection with sonographicimaging. An echogenic agent can, in some embodiments, be used to enhancethe sonographic signal, providing targeting for the imaging, or performother functions, as will be recognized by one of ordinary skill in theart. As used herein, the term “glycomimetic” is used as conventional inthe art and refers to a compound that mimics the function of a bioactivecarbohydrate.

It should be noted that as used herein and in the appended claims, thesingular forms “a”, “an”, and “the” include plural reference unless thecontext clearly dictates otherwise. Thus, for example, reference to “acell” includes a plurality of such cells and equivalents thereof knownto those skilled in the art, and so forth. As well, the terms “a” (or“an”), “one or more” and “at least one” can be used interchangeablyherein. It is also to be noted that the terms “comprising”, “including”,and “having” can be used interchangeably.

In certain embodiments, the invention encompasses administering opticalagents useful in the invention to a patient or subject. A “patient” or“subject”, used equivalently herein, refers to an animal. In particular,an animal refers to a mammal, preferably a human. The subject mayeither: (1) have a condition diagnosable, preventable and/or treatableby administration of an optical agent of the invention; or (2) issusceptible to a condition that is diagnosable, preventable and/ortreatable by administering an optical agent of this invention. Thecompounds of the invention function as optical agents, in someembodiments.

As to any of the above groups which contain one or more substituents, itis understood, that such groups do not contain any substitution orsubstitution patterns which are sterically impractical and/orsynthetically non-feasible. In addition, the compounds of this inventioninclude all stereochemical isomers arising from the substitution ofthese compounds.

The compounds of this invention may contain one or more chiral centers.Accordingly, this invention is intended to include racemic mixtures,diasteromers, enantiomers and mixture enriched in one or moresteroisomer. The scope of the invention as described and claimedencompasses the racemic forms of the compounds as well as the individualenantiomers and non-racemic mixtures thereof.

The invention is further detailed in the following Examples, which areoffered by way of illustration and are not intended to limit the scopeof the invention in any manner.

Example 1: Formulations of Non-Benzenoid Aromatic Compounds

In an embodiment, the invention provides a pharmaceutical formulationcomprising a composition of the invention, such as a compound of any oneof formulae (FX1)-(FX17). In an embodiment, the invention provides amethod of synthesizing a composition of the invention or apharmaceutical formulation thereof, such as a compound of any one offormulae (FX1)-(FX17). In an embodiment, the invention provides apharmaceutical formulation comprising a composition of the invention,such as a compound of formula (FX18). In an embodiment, the inventionprovides a method of synthesizing a composition of the invention or apharmaceutical formulation thereof, such as a compound of formula(FX18). In an embodiment, a pharmaceutical formulation comprises one ormore excipients, carriers, diluents, and/or other components as would beunderstood in the art. Preferably, the components meet the standards ofthe National Formulary (“NF”), United States Pharmacopoeia (“USP”;United States Pharmacopeia Convention Inc., Rockville, Md.), or Handbookof Pharmaceutical Manufacturing Formulations (Sarfaraz K. Niazi, allvolumes, ISBN: 9780849317521, ISBN 10: 0849317525; CRC Press, 2004).See, e.g., United States Pharmacopeia and National Formulary (USP 30-NF25), Rockville, Md.: United States Pharmacopeial Convention; 2007; and2008, and each of any earlier editions; The Handbook of PharmaceuticalExcipients, published jointly by the American Pharmacists Associationand the Pharmaceutical Press (Pharmaceutical Press (2005) (ISBN-10:0853696187, ISBN-13: 978-0853696186); Merck Index, Merck & Co., Rahway,N.J.; and Gilman et al., (eds) (1996); Goodman and Gilman's: ThePharmacological Bases of Therapeutics, 8th Ed., Pergamon Press. Inembodiments, the formulation base of the formulations of the inventioncomprises physiologically acceptable excipients, namely, at least onebinder and optionally other physiologically acceptable excipients.Physiologically acceptable excipients are those known to be usable inthe pharmaceutical technology sectors and adjacent areas, particularly,those listed in relevant pharmacopeias (e.g. DAB, Ph. Eur., BP, NF,USP), as well as other excipients whose properties do not impair aphysiological use.

In an embodiment, an effective amount of a composition of the inventionis a therapeutically effective amount. In an embodiment, an effectiveamount of a composition of the invention is a diagnostically effectiveamount. In an embodiment, an active ingredient or other component isincluded in a therapeutically acceptable amount. In an embodiment, anactive ingredient or other component is included in a diagnosticallyacceptable amount.

Variations on compositions including salts and ester forms of compounds:Compounds of this invention and compounds useful in the methods of thisinvention include those of the compounds and formula(s) described hereinand pharmaceutically-acceptable salts and esters of those compounds. Inembodiments, salts include any salts derived from the acids of theformulas herein which acceptable for use in human or veterinaryapplications. In embodiments, the term “esters” refers to hydrolyzableesters of compounds of the names and structural formulas herein. Inembodiments, salts and esters of the compounds of the formulas hereincan include those which have the same or better therapeutic, diagnostic,or pharmaceutical (human or veterinary) general properties as thecompounds of the formulas herein. In an embodiment, a composition of theinvention is a compound or salt or ester thereof suitable forpharmaceutical formulations.

In an embodiment, the invention provides a method for treating a medicalcondition comprising administering to a subject (e.g. patient) in needthereof, a therapeutically effective amount of a composition of theinvention, such as a compound of any one of formulae (FX1)-(FX17). In anembodiment, the invention provides a method for treating a medicalcondition comprising administering to a subject (e.g. patient) in needthereof, a therapeutically effective amount of a composition of theinvention, such as a compound of formula (FX18). In an embodiment, themedical condition is cancer, or various other diseases, injuries, anddisorders, including cardiovascular disorders such as atherosclerosisand vascular restenosis, inflammatory diseases, ophthalmic diseases anddermatological diseases.

In an embodiment, the invention provides a medicament which comprises atherapeutically effective amount of one or more compositions of theinvention, such as a compound of any one of formulae (FX1)-(FX17). In anembodiment, the invention provides a medicament which comprises atherapeutically effective amount of one or more compositions of theinvention, such as a compound of formula (FX18). In an embodiment, theinvention provides a medicament which comprises a therapeutically ordiagnostically effective amount of one or more compositions of theinvention. In an embodiment, the invention provides a method for makinga medicament for treatment of a condition described herein. In anembodiment, the invention provides a method for making a medicament fordiagnosis or aiding in the diagnosis of a condition described herein. Inan embodiment, the invention provides the use of one or morecompositions set forth herein for the making of a medicament. In anembodiment, the invention provides one or more compositions set forthherein for use as a medicament. In an embodiment, the invention providesone or more compositions set forth herein for use in therapy.

Compounds of the invention can have prodrug forms. Prodrugs of thecompounds of the invention are useful in embodiments includingcompositions and methods. Any compound that will be converted in vivo toprovide a biologically, pharmaceutically, diagnostically, ortherapeutically active form of a compound of the invention is a prodrug.Various examples and forms of prodrugs are well known in the art.Examples of prodrugs are found, inter alia, in Design of Prodrugs,edited by H. Bundgaard, (Elsevier, 1985), Methods in Enzymology, Vol.42, at pp. 309-396, edited by K. Widder, et. al. (Academic Press, 1985);A Textbook of Drug Design and Development, edited by Krosgaard-Larsenand H. Bundgaard, Chapter 5, “Design and Application of Prodrugs,” by H.Bundgaard, at pp. 113-191, 1991); H. Bundgaard, Advanced Drug DeliveryReviews, Vol. 8, p. 1-38 (1992); H. Bundgaard, et al., Journal ofPharmaceutical Sciences, Vol. 77, p. 285 (1988); and Nogrady (1985)Medicinal Chemistry A Biochemical Approach, Oxford University Press, NewYork, pages 388-392). A prodrug, such as a pharmaceutically acceptableprodrug can represent prodrugs of the compounds of the invention whichare, within the scope of sound medical judgment, suitable for use incontact with the tissues of humans and lower animals without unduetoxicity, irritation, allergic response, and the like, commensurate witha reasonable benefit/risk ratio, and effective for their intended use.Prodrugs of the invention can be rapidly transformed in vivo to a parentcompound of a compound described herein, for example, by hydrolysis inblood or by other cell, tissue, organ, or system processes. Furtherdiscussion is provided in T. Higuchi and V. Stella, Pro-drugs as NovelDelivery Systems, V. 14 of the A.C.S. Symposium Series, and in Edward B.Roche, ed., Bioreversible Carriers in Drug Design, AmericanPharmaceutical Association and Pergamon Press (1987).

The invention contemplates pharmaceutically active compounds eitherchemically synthesized or formed by in vivo biotransformation tocompounds set forth herein.

In an embodiment, a composition of the invention is isolated orpurified. In an embodiment, an isolated or purified compound may be atleast partially isolated or purified as would be understood in the art.In an embodiment, the composition of the invention has a chemical purityof 95%, optionally for some applications 99%, optionally for someapplications 99.9%, optionally for some applications 99.99%, andoptionally for some applications 99.999% pure.

Typically, a compound of the present invention, or pharmaceuticallyacceptable salt thereof, is administered to a subject in adiagnostically or therapeutically effective amount. One skilled in theart generally can determine an appropriate dosage. Factors affecting aparticular dosage regimen (including the amount of compound delivered,frequency of administration, and whether administration is continuous orintermittent) include, for example, the type, age, weight, sex, diet,and condition of the subject; the type of pathological condition and itsseverity; and the nature of the desired effect. Pharmacologicalconsiderations include non-benzenoid compound activity, efficacy,pharmacokinetic, and toxicology profiles of the particular non-benzenoidcompound used; the route of administration and whether a drug deliverysystem is utilized; and whether the non-benzenoid compound isadministered as part of a combination therapy (e.g., whether the agentis administered in combination with one or more active compounds, otheragents, radiation, and the like).

Compositions for oral administration may be, for example, prepared in amanner such that a single dose in one or more oral preparations containsat least about 20 mg of the non-benzenoid compound per square meter ofsubject body surface area, or at least about 50, 100, 150, 200, 300,400, or 500 mg of the non-benzenoid compound per square meter of subjectbody surface area (the average body surface area for a human is, forexample, 1.8 square meters). In particular, a single dose of acomposition for oral administration can contain from about 20 to about600 mg, and in certain aspects from about 20 to about 400 mg, in anotheraspect from about 20 to about 300 mg, and in yet another aspect fromabout 20 to about 200 mg of the non-benzenoid compound per square meterof subject body surface area. Compositions for parenteral administrationcan be prepared in a manner such that a single dose contains at leastabout 20 mg of the non-benzenoid compound per square meter of subjectbody surface area, or at least about 40, 50, 100, 150, 200, 300, 400, or500 mg of the non-benzenoid compound per square meter of subject bodysurface area. In particular, a single dose in one or more parenteralpreparations contains from about 20 to about 500 mg, and in certainaspects from about 20 to about 400, and in another aspect from about 20to about 400 mg, and in yet another aspect from about 20 to about 350 mgof the non-benzenoid compound per square meter of subject body surfacearea. It should be recognized that these oral and parenteral dosageranges represent generally preferred dosage ranges, and are not intendedto limit the invention. The dosage regimen actually employed can varywidely, and, therefore, can deviate from the generally preferred dosageregimen. It is contemplated that one skilled in the art will tailorthese ranges to the individual subject.

As indicated above, it is contemplated that the compounds andpharmaceutically acceptable salts of the present invention may be usedas part of a combination. The term “combination” means theadministration of two or more compounds directed to the targetcondition. The treatments of the combination generally may beco-administered in a simultaneous manner. Two compounds can beco-administered as, for example: (a) a single formulation (e.g., asingle capsule) having a fixed ratio of active ingredients; or (b)multiple, separate formulations (e.g., multiple capsules) for eachcompound. The treatments of the combination may alternatively (oradditionally) be administered at different times.

It is further contemplated that the non-benzenoid compounds and salts ofthis invention can be used in the form of a kit that is suitable for usein performing the methods described herein, packaged in a container. Thekit can contain the non-benzenoid compound or compounds and, optionally,appropriate diluents, devices or device components suitable foradministration and instructions for use in accordance with the methodsof the present invention. The devices can include parenteral injectiondevices, such as syringes or transdermal patch or the like. Devicecomponents can include cartridges for use in injection devices and thelike. In one aspect, the kit includes a first dosage form including anon-benzenoid compound or salt of this invention and a second dosageform including another active ingredient in quantities sufficient tocarry out the methods of the present invention. The first dosage formand the second dosage form together can include a therapeuticallyeffective amount of the compounds for treating the targetedcondition(s).

This invention also is directed, in part, to pharmaceutical compositionsincluding a therapeutically effective amount of a compound or salt ofthis invention, as well as processes for making such compositions. Suchcompositions generally include one or more pharmaceutically acceptablecarriers (e.g., excipients, vehicles, auxiliaries, adjuvants, diluents)and may include other active ingredients. Formulation of thesecompositions may be achieved by various methods known in the art. Ageneral discussion of these methods may be found in, for example,Hoover, John E., Remington's Pharmaceutical Sciences (Mack PublishingCo., Easton, Pa.: 1975). See also, Lachman, L., eds., PharmaceuticalDosage Forms (Marcel Decker, New York, N.Y., 1980).

The preferred composition depends on the route of administration. Anyroute of administration may be used as long as the target of thecompound or pharmaceutically acceptable salt is available via thatroute. Suitable routes of administration include, for example, oral,parenteral, inhalation, rectal, nasal, topical (e.g., transdermal andintraocular), intravesical, intrathecal, enteral, pulmonary,intralymphatic, intracavital, vaginal, transurethral, intradermal,aural, intramammary, buccal, orthotopic, intratracheal, intralesional,percutaneous, endoscopical, transmucosal, sublingual, and intestinaladministration.

Pharmaceutically acceptable carriers that may be used in conjunctionwith the compounds of the invention are well known to those of ordinaryskill in the art. Carriers can be selected based on a number of factorsincluding, for example, the particular non-benzenoid compound(s) orpharmaceutically acceptable salt(s) used; the compound's concentration,stability, and intended bioavailability; the condition being treated;the subject's age, size, and general condition; the route ofadministration; etc. A general discussion related to carriers may befound in, for example, J. G. Nairn, Remington's Pharmaceutical Science,pp. 1492-1517 (A. Gennaro, ed., Mack Publishing Co., Easton, Pa.(1985)).

Solid dosage forms for oral administration include, for example,capsules, tablets, gelcaps, pills, dragees, troches, powders, granules,and lozenges. In such solid dosage forms, the compounds orpharmaceutically acceptable salts thereof can be combined with one ormore pharmaceutically acceptable carriers. The compounds andpharmaceutically acceptable salts thereof can be mixed with carriersincluding, but not limited to, lactose, sucrose, starch powder, cornstarch, potato starch, magnesium carbonate, microcrystalline cellulose,cellulose esters of alkanoic acids, cellulose alkyl esters, talc,stearic acid, magnesium stearate, magnesium oxide, sodium and calciumsalts of phosphoric and sulfuric acids, sodium carbonate, agar,mannitol, sorbitol, sodium saccharin, gelatin, acacia gum, alginic acid,sodium alginate, tragacanth, colloidal silicon dioxide, croscarmellosesodium, polyvinylpyrrolidone, and/or polyvinyl alcohol, and thentableted or encapsulated for convenient administration. Such capsules ortablets can contain a controlled-release formulation, as can be providedin a dispersion of the compound or salt in hydroxypropylmethylcellulose. In the case of capsules, tablets, and pills, the dosage formsalso can include buffering agents, such as sodium citrate, or magnesiumor calcium carbonate or bicarbonate. Tablets and pills additionally can,for example, include a coating (e.g., an enteric coating) to delaydisintegration and absorption. The concentration of the non-benzenoidcompound in a solid oral dosage form can be from about 5 to about 50%,and in certain aspects from about 8 to about 40%, and in another aspectfrom about 10 to about 30% by weight based on the total weight of thecomposition.

Liquid dosage forms of the compounds of the present invention for oraladministration include, for example, pharmaceutically acceptableemulsions, solutions, suspensions, syrups, and elixirs containing inertdiluents commonly used in the art (e.g., water). Such compositions alsocan include adjuvants, such as wetting, emulsifying, suspending,flavoring (e.g., sweetening), and/or perfuming agents. The concentrationof the non-benzenoid compound in the liquid dosage form can be fromabout 0.01 to about 5 mg, and in certain aspects from about 0.01 toabout 1 mg, and in another aspect from about 0.01 to about 0.5 mg per mlof the composition. Low concentrations of the compounds of the presentinvention in liquid dosage form can be prepared in the case that thenon-benzenoid compound is more soluble at low concentrations. Techniquesfor making oral dosage forms useful in the present invention aregenerally described in, for example, Modern Pharmaceutics, Chapters 9and 10 (Banker & Rhodes, Editors (1979)). See also, Lieberman et al.,Pharmaceutical Dosage Forms: Tablets (1981). See also, Ansel,Introduction to Pharmaceutical Dosage Forms (2nd Edition (1976)).

In some aspects of the present invention, tablets or powders for oraladministration can be prepared by dissolving the non-benzenoid compoundin a pharmaceutically acceptable solvent capable of dissolving thecompound to form a solution and then evaporating when the solution isdried under vacuum. A carrier can also be added to the solution beforedrying. The resulting solution can be dried under vacuum to form aglass. The glass can then mix with a binder to form a powder. Thispowder may be mixed with fillers or other conventional tableting agents,and then processed to form a tablet. Alternatively, the powder may beadded to a liquid carrier to form a solution, emulsion, suspension, orthe like.

In some aspects, solutions for oral administration are prepared bydissolving the non-benzenoid compound in a pharmaceutically acceptablesolvent capable of dissolving the compound to form a solution. Anappropriate volume of a carrier is added to the solution while stirringto form a pharmaceutically acceptable solution for oral administration.

“Parenteral administration” includes subcutaneous injections,intravenous injections, intraarterial injections, intraorbitalinjections, intracapsular injections, intraspinal injections,intraperitoneal injections, intramuscular injections, intrasternalinjections, and infusion. Dosage forms suitable for parenteraladministration include solutions, suspensions, dispersions, emulsions,and any other dosage form that can be administered parenterally.

Injectable preparations (e.g., sterile injectable aqueous or oleaginoussuspensions) can be formulated according to the known art using suitabledispersing, wetting agents, and/or suspending agents. Acceptablevehicles for parenteral use include both aqueous and nonaqueouspharmaceutically-acceptable solvents. Suitable pharmaceuticallyacceptable aqueous solvents include, for example, water, salinesolutions, dextrose solutions (e.g., such as DW5), electrolytesolutions, etc.

Suitable pharmaceutically-acceptable nonaqueous solvents include, butare not limited to, the following (as well as mixtures thereof):alcohols (these include, for example, σ-glycerol formal, β-glycerolformal, 1,3-butyleneglycol, aliphatic or aromatic alcohols having from 2to about 30 carbons (e.g., methanol, ethanol, propanol, isopropanol,butanol, t-butanol, hexanol, octanol, amylene hydrate, benzyl alcohol,glycerin (glycerol), glycol, hexylene, glycol, tetrahydrofuranylalcohol, cetyl alcohol, and stearyl alcohol), fatty acid esters of fattyalcohols (e.g., polyalkylene glycols, such as polypropylene glycol andpolyethylene glycol), sorbitan, sucrose, and cholesterol); amides (theseinclude, for example, dimethylacetamide (DMA), benzyl benzoate DMA,dimethylformamide, N-hydroxyethyO-lactamide, N,N-dimethylacetamide-amides, 2-pyrrolidinone, 1-methyl-2-pyrrolidinone,and polyvinyl pyrrolidone); esters (these include, for example, acetateesters (e.g., monoacetin, diacetin, and triacetin), aliphatic andaromatic esters (e.g., ethyl caprylate or octanoate, alkyl oleate,benzyl benzoate, or benzyl acetate), dimethylsulfoxide (DMSO), esters ofglycerin (e.g., mono, di, and tri-glyceryl citrates and tartrates),ethyl benzoate, ethyl acetate, ethyl carbonate, ethyl lactate, ethyloleate, fatty acid esters of sorbitan, glyceryl monostearate, glycerideesters (e.g., mono, di, or tri-glycerides), fatty acid esters (e.g.,isopropyl myristrate), fatty acid derived PEG esters (e.g.,PEG-hydroxyoleate and PEG-hydroxystearate), N-methyl pyrrolidinone,pluronic 60, polyoxyethylene sorbitol oleic polyesters (e.g.,poly(ethoxylated)₃₀₋₆₀ sorbitol poly(oleate)₂₋₄, poly(oxyethylene)₁₅₋₂₀monooleate, poly(oxyethylene)₁₅₋₂₀ mono 12-hydroxystearate, andpoly(oxyethylene)₁₅₋₂₀ mono ricinoleate), polyoxyethylene sorbitanesters (e.g., polyoxyethylene-sorbitan monooleate,polyoxyethylene-sorbitan monopalmitate, polyoxyethylene-sorbitanmonolaurate, polyoxyethylene-sorbitan monostearate, and POLYSORBATE 20,40, 60, and 80 (from ICI Americas, Wilmington, Del.)),polyvinylpyrrolidone, alkyleneoxy modified fatty acid esters (e.g.,polyoxyl 40 hydrogenated castor oil and polyoxyethylated castor oils,such as CREMOPHOR EL solution or CREMOPHOR RH 40 solution), saccharidefatty acid esters (i.e., the condensation product of a monosaccharide(e.g., pentoses, such as, ribose, ribulose, arabinose, xylose, lyxose,and xylulose; hexoses, such as glucose, fructose, galactose, mannose,and sorbose; trioses; tetroses; heptoses; and octoses), disaccharide(e.g., sucrose, maltose, lactose, and trehalose), oligosaccharide, or amixture thereof with one or more C₄-C₂₂ fatty acids (e.g., saturatedfatty acids, such as caprylic acid, capric acid, lauric acid, myristicacid, palmitic acid, and stearic acid; and unsaturated fatty acids, suchas palmitoleic acid, oleic acid, elaidic acid, erucic acid, and linoleicacid), and steroidal esters); ethers (these are typically alkyl, aryl,and cyclic ethers having from 2 to about 30 carbons. Examples includediethyl ether, tetrahydrofuran, dimethyl isosorbide, diethylene glycolmonoethyl ether), and glycofurol (tetrahydrofurfuranyl alcoholpolyethylene glycol ether); ketones (these typically have from about 3to about 30 carbons. Examples include acetone, methyl ethyl ketone,methyl isobutyl ketone); hydrocarbons (these are typically aliphatic,cycloaliphatic, and aromatic hydrocarbons having from about 4 to about30 carbons). Examples include benzene, cyclohexane, dichloromethane,dioxolanes, hexane, n-decane, n-dodecane, n-hexane, sulfolane,tetramethylenesulfone, tetramethylenesulfoxide, toluene,dimethylsulfoxide (DMSO); and tetramethylene sulfoxide; oils (theseinclude oils of mineral, vegetable, animal, essential, or syntheticorigin). These include mineral oils, such as aliphatic and wax-basedhydrocarbons, aromatic hydrocarbons, mixed aliphatic and aromatic basedhydrocarbons, and refined paraffin oil; vegetable oils, such as linseed,tung, safflower, soybean, castor, cottonseed, groundnut, rapeseed,coconut, palm, olive, corn, corn germ, sesame, persic, and peanut oil;glycerides, such as mono-, di-, and triglycerides; animal oils, such asfish, marine, sperm, cod-liver, haliver, squaiene, squalane, and sharkliver oil; oleic oils; and polyoxyethylated castor oil); alkyl, alkenyl,or aryl halides (these include alkyl or aryl halides having from 1 toabout 30 carbons and one or more halogen substituents. Examples includemethylene chloride); monoethanolamine; petroleum benzin; trolamine;omega-3 polyunsaturated fatty acids (e.g., alpha-linolenic acid,eicosapentaenoic acid, docosapentaenoic acid, or docosahexaenoic acid);polyglycol ester of 12-hydroxystearic acid and polyethylene glycol(SOLUTOL HS-15, from BASF, Ludwigshafen, Germany); polyoxyethyleneglycerol; sodium laurate; sodium oleate; and sorbitan monooleate. Otherpharmaceutically acceptable solvents for use in the invention are wellknown to those of ordinary skill in the art. General discussion relatingto such solvents may be found in, for example, The Chemotherapy SourceBook (Williams & Wilkens Publishing), The Handbook of PharmaceuticalExcipients, (American Pharmaceutical Association, Washington, D.C., andThe Pharmaceutical Society of Great Britain, London, England, 1968),Modern Pharmaceutics 3d ed., (G. Banker et. al., eds., Marcel Dekker,Inc., New York, N.Y. (1995)), The Pharmacological Basis of Therapeutics,(Goodman & Gilman, McGraw Hill Publishing), Pharmaceutical Dosage Forms,(H. Lieberman et. al., eds., Marcel Dekker, Inc., New York, N.Y.(1980)), Remington's Pharmaceutical Sciences, 19th ed., (A. Gennaro,ed., Mack Publishing, Easton, Pa., (1995)), The United StatesPharmacopeia 24, The National Formulary 19, (National Publishing,Philadelphia, Pa. (2000)); Spiegel, A. J., et al., “Use of NonaqueousSolvents in Parenteral Products,” J. Pharma. Sciences, Vol. 52, No. 10,pp. 917-927 (1963).

Solvents useful in the present invention include, but are not limitedto, those known to stabilize the non-benzenoid compounds orpharmaceutically acceptable salts thereof. These typically include, forexample, oils rich in triglycerides, such as safflower oil, soybean oil,and mixtures thereof; and alkyleneoxy-modified fatty acid esters, suchas polyoxyl 40 hydrogenated castor oil and polyoxyethylated castor oils(e.g., CREMOPHOR EL solution or CREMOPHOR RH 40 solution). Commerciallyavailable triglycerides include INTRALIPID emulsified soybean oil(Kabi-Pharmacia Inc., Stockholm, Sweden), NUTRALIPID emulsion (McGaw,Irvine, Calif.), LIPOSYN II 20% emulsion (a 20% fat emulsion solutioncontaining 100 mg safflower oil, 100 mg soybean oil, 12 mg eggphosphatides, and 25 mg glycerin per ml of solution; AbbottLaboratories, Chicago, Ill.), LIPOSYN III 2% emulsion (a 2% fat emulsionsolution containing 100 mg safflower oil, 100 mg soybean oil, 12 mg eggphosphatides, and 25 mg glycerin per ml of solution; AbbottLaboratories, Chicago, Ill.), natural or synthetic glycerol derivativescontaining the docosahexaenoyl group at levels of from about 25 to about100% (by weight based on the total fatty acid content) (DHASCO fromMartek Biosciences Corp., Columbia, Md.; DHA MAGURO from DaitoEnterprises, Los Angeles, Calif.; SOYACAL; and TRAVEMULSION). Ethanol inparticular is a useful solvent for dissolving a non-benzenoid compoundor pharmaceutically acceptable salt thereof to form solutions,emulsions, and the like.

Additional components can be included in the compositions of thisinvention for various purposes generally known in the pharmaceuticalindustry. These components tend to impart properties that, for example,enhance retention of the non-benzenoid compound or salt at the site ofadministration, protect the stability of the composition, control thepH, and facilitate processing of the non-benzenoid compound or salt intopharmaceutical formulations, and the like. Specific examples of suchcomponents include cryoprotective agents; agents for preventingreprecipitation of the non-benzenoid compound or salt surface; active,wetting, or emulsifying agents (e.g., lecithin, polysorbate-80, TWEEN80, pluronic 60, and polyoxyethylene stearate); preservatives (e.g.,ethyl-p-hydroxybenzoate); microbial preservatives (e.g., benzyl alcohol,phenol, m-cresol, chlorobutanol, sorbic acid, thimerosal, and paraben);agents for adjusting pH or buffering agents (e.g., acids, bases, sodiumacetate, sorbitan monolaurate, etc.); agents for adjusting osmolarity(e.g., glycerin); thickeners (e.g., aluminum monostearate, stearic acid,cetyl alcohol, stearyl alcohol, guar gum, methyl cellulose,hydroxypropylcellulose, tristearin, cetyl wax esters, polyethyleneglycol, etc.); colorants; dyes; flow aids; non-volatile silicones (e.g.,cyclomethicone); clays (e.g., bentonites); adhesives; bulking agents;flavorings; sweeteners; adsorbents; fillers (e.g., sugars such aslactose, sucrose, mannitol, sorbitol, cellulose, calcium phosphate,etc.); diluents (e.g., water, saline, electrolyte solutions, etc.);binders (e.g., gelatin; gum tragacanth; methyl cellulose; hydroxypropylmethylcellulose; sodium carboxymethyl cellulose; polyvinylpyrrolidone;sugars; polymers; acacia; starches, such as maize starch, wheat starch,rice starch, and potato starch; etc.); disintegrating agents (e.g.,starches, such as maize starch, wheat starch, rice starch, potatostarch, and carboxymethyl starch; cross-linked polyvinyl pyrrolidone;agar; alginic acid or a salt thereof, such as sodium alginate;croscarmellose sodium; crospovidone; etc); lubricants (e.g., silica;talc; stearic acid and salts thereof, such as magnesium stearate;polyethylene glycol; etc.); coating agents (e.g., concentrated sugarsolutions including gum arabic, talc, polyvinyl pyrrolidone, carbopolgel, polyethylene glycol, titanium dioxide, etc.); and antioxidants(e.g., sodium metabisulfite, sodium bisulfite, sodium sulfite, dextrose,phenols, thiophenols, etc.). Techniques and compositions for makingparenteral dosage forms are generally known in the art. Formulations forparenteral administration may be prepared from one or more sterilepowders and/or granules having a compound or salt of this invention andone or more of the carriers or diluents mentioned for use in theformulations for oral administration. The powder or granule typically isadded to an appropriate volume of a solvent (typically while agitating(e.g., stirring) the solvent) that is capable of dissolving the powderor granule. Particular solvents useful in the invention include, forexample, water, polyethylene glycol, propylene glycol, ethanol, cornoil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodiumchloride, and/or various buffers.

Emulsions for parenteral administration can be prepared by, for example,dissolving a compound or salt of this invention in any pharmaceuticallyacceptable solvent capable of dissolving the compound to form asolution; and adding an appropriate volume of a carrier, which is anemulsion, to the solution while stirring to form the emulsion. Solutionsfor parenteral administration can be prepared by, for example,dissolving a compound or salt of this invention in any pharmaceuticallyacceptable solvent capable of dissolving the compound to form asolution; and adding an appropriate volume of a carrier to the solutionwhile stirring to form the solution.

Suppositories for rectal administration can be prepared by, for example,mixing the drug with a suitable nonirritating excipient that is solid atordinary temperatures, but liquid at the rectal temperature and willtherefore melt in the rectum to release the drug. Suitable excipientsinclude, for example, cocoa butter; synthetic mono-, di-, ortriglycerides; fatty acids; and/or polyethylene glycols.

“Topical administration” includes the use of transdermal administration,such as transdermal patches or iontophoresis devices.

If desired, the emulsions or solutions described above for oral orparenteral administration can be packaged in IV bags, vials, or otherconventional containers in concentrated form, and then diluted with apharmaceutically acceptable liquid (e.g., saline) to form an acceptablenon-benzenoid concentration before use.

Other adjuvants and modes of administration well known in thepharmaceutical art may also be used. Pharmaceutically acceptable saltscomprise pharmaceutically-acceptable anions and/or cations.Pharmaceutically-acceptable cations include among others, alkali metalcations (e.g., Li⁺, Na⁺, K⁺), alkaline earth metal cations (e.g., Ca²⁺,Mg²⁺), non-toxic heavy metal cations and ammonium (NH₄ ⁺) andsubstituted ammonium (N(R′)₄ ⁺, where R′ is hydrogen, alkyl, orsubstituted alkyl, i.e., including, methyl, ethyl, or hydroxyethyl,specifically, trimethyl ammonium, triethyl ammonium, and triethanolammonium cations). Pharmaceutically-acceptable anions include amongother halides (e.g., Cl⁻, Br⁻), sulfate, acetates (e.g., acetate,trifluoroacetate), ascorbates, aspartates, benzoates, citrates, andlactate.

It is understood that this invention is not limited to the particularcompounds, methodology, protocols, and reagents described, as these mayvary. It is also to be understood that the terminology used herein isfor the purpose of describing particular embodiments only, and is notintended to limit the scope of the invention which will be limited onlyby the appended claims.

Compositions of the invention includes formulations and preparationscomprising one or more of the present compounds provided in an aqueoussolution, such as a pharmaceutically acceptable formulation orpreparation. Optionally, compositions of the invention further compriseone or more pharmaceutically acceptable surfactants, buffers,electrolytes, salts, carriers, binders, coatings, preservatives and/orexcipients.

Example 2: Optical Imaging Using Non-Benzenoid Compounds

In general, molecules absorbing, emitting, or scattering in the visibleor NIR region of the electromagnetic spectrum are useful for opticalmeasurement. The high sensitivity associated with fluorescence permitsdetection without the negative effects of radioactivity or ionizingradiation. Non-benzenoid aromatic compounds of the invention absorbstrongly in the red and NIR regions. Furthermore, the electronicproperties of these systems are very sensitive to substitution patternsin both rings of the non-benzenoid compound and allows for “tuning” theabsorption and emission properties using the information describedherein.

In an embodiment of this aspect, the invention provides a method ofusing an optical agent, for example, in a biomedical procedure foroptically imaging or visualizing a target tissue or a class of targettissues. The present methods include tissue selective imaging andvisualization methods, such as imaging or visualization of renal tissue.A method of this aspect comprises the step of administering adiagnostically effective amount of a compound to a subject, wherein thecompound is a compound having any of formulae (FX1) to (FX17) or apharmaceutical preparation thereof. A method of this aspect comprisesthe step of administering a diagnostically effective amount of acompound to a subject, wherein the compound is a compound having formula(FX18) or a pharmaceutical preparation thereof. The present methods areuseful for imaging or visualizing colorectal cancer and other cancers,including prostate cancer, gastric cancer, esophageal cancer,uterine-endometrial cancer, pancreatic cancer, breast cancer, cervicalcancer, head and neck cancer, hepatic cancer, skin cancer, gallbladdercancer, ling cancer and ovarian cancer.

In methods of this aspect, the compound that has been administered tothe subject then is exposed in vivo to electromagnetic radiation andelectromagnetic radiation emitted or scattered by the compound is thendetected. In some embodiments, fluorescence is excited from the compound(e.g., due to the electromagnetic radiation exposure), optionally viamultiphoton excitation processes. In an embodiment particularly usefulfor imaging and/or visualization, the method of this aspect furthercomprises: (i) exposing a compound, such as a compound having any one offormula (FX1) to (FX17), administered to the subject to electromagneticradiation capable of exciting emission from the compound; and (ii)measuring the emission from the compound. In an embodiment particularlyuseful for imaging and/or visualization, the method of this aspectfurther comprises: (i) exposing a compound, such as a compound havingformula (FX18), administered to the subject to electromagnetic radiationcapable of exciting emission from the compound; and (ii) measuring theemission from the compound. In some embodiments, the methods of thepresent invention use fluorescence excitation via exposure to lighthaving wavelengths selected over the range of 400-1300 nm. For example,optical coherence tomography (OCT) is an optical imaging techniquecompatible with the present compounds that allows high resolution crosssectional imaging of tissue microstructure. OCT methods use wavelengthsof about 1280 nm. Use of electromagnetic radiation having wavelengthsselected over the range of 700 nanometers to 1300 nanometers may beuseful for some in situ optical imaging methods of the presentinvention, including biomedical applications for imaging organs, tissueand/or tumors, anatomical visualization, optical guided surgery andendoscopic procedures. Compounds in present methods may function ascontrast agents, optical probes and/or tracer elements. The methods ofthe present invention include in vivo, in vitro and ex vivo imaging andvisualization. The present invention provides methods for a range ofclinical procedures, including optical imaging methods and/orvisualization guided surgery and/or endoscopic diagnostic andtherapeutic procedures.

In an exemplary protocol of uses of the compounds of the invention for abiomedical imaging procedure, the non-benzenoid aromatic compound isexposed to visible and/or near infrared light. This exposure of thenon-benzenoid aromatic compound to light may occur at any appropriatetime but preferably occurs while the non-benzenoid aromatic compound islocated in the body. Due to this exposure of the non-benzenoid aromaticcompound to the visible and/or infrared light, the non-benzenoidderivative emits spectral energy (e.g., visible and/or near infraredlight) that may be detected by appropriate detection equipment. Thespectral energy emitted from the non-benzenoid aromatic compound tendsto exhibit a wavelength range greater than a wavelength range absorbedby the non-benzenoid aromatic compound. For example, if thenon-benzenoid aromatic compound absorbs light of about 700 nm, thenon-benzenoid aromatic compound may emit light of about 745 nm.

Detection of the non-benzenoid aromatic compound (or more particularly,light emitted therefrom) may be achieved through optical fluorescence,absorbance or light scattering procedures known in the art. Thisdetection of the emitted spectral energy, or luminescence, may becharacterized as a collection of the emitted spectral energy and ageneration of electrical signal indicative of the collected spectralenergy. For these purposes, the term “luminescence” refers to theemission of light from excited electronic states of atoms or molecules.Luminescence generally refers to light emission, such asphotoluminescence, chemiluminescence, and electrochemiluminescence,among others. In photoluminescence, including fluorescence andphosphorescence, the excited electronic state is created by theabsorption of electromagnetic radiation. Luminescence detection involvesdetection of one or more properties of the luminescence or associatedluminescence process. These properties may include intensity, excitationand/or emission spectrum, polarization, lifetime, and energy transfer,among others. These properties may also include time-independent(steady-state) and/or time-dependent (time-resolved) properties of theluminescence. Representative luminescence techniques includefluorescence intensity (FLINT), fluorescence polarization (FP),fluorescence resonance energy transfer (FRET), fluorescence lifetime(FLT), total internal reflection fluorescence (TIRF), fluorescencecorrelation spectroscopy (FCS), fluorescence recovery afterphotobleaching (FRAP), optical-acoustic tomography (OAT) andbioluminescence resonance energy transfer (BRET), multiphotontechnology, among others.

By way of example, when a compound is used in the present invention, itis desirable that the wavelength of light supplied to the compound besuch that it excites the compound. This excitation causes the moleculeto emit part of the absorbed energy at a different wavelength, and theemission can be detected using fluorometric techniques or othertechniques as described above. One skilled in the art can readilydetermine the most appropriate detection technique based on, in part,the specific compound(s) administered, the particular use (e.g., tissueto be detected) and other aspects, including physical limitations of theanalysis.

The techniques utilized to detect the spectral energy from thenon-benzenoid derivative that is present in the body may be designed todetect only selected wavelengths (or wavelength ranges) and/or mayinclude one or more appropriate spectral filters. Various catheters,endoscopes, ear clips, headbands, surface coils, finger probes, and thelike may be utilized to expose the non-benzenoid derivative to lightand/or to detect light emitting therefrom. This detection of spectralenergy may be accomplished at one or more times intermittently or may besubstantially continuous.

Preferably, non-ionizing energy is administered to the subject or samplefor detecting or imaging a biological sample to a compound of theinvention. For these purposes, the term “non-ionizing energy” generallyrefers to electromagnetic radiation that does not carry enough energy tocompletely remove at least one electron from an atom or molecule of thepatient's body. For example, in some embodiments, non-ionizing energymay include spectral energy ranging in wavelength from about 400 nm toabout 1200 nm. In some embodiments, non-ionizing energy may simplyinclude visible and/or near infrared light.

In one embodiment, the spectral properties of the compounds of theinvention may be tuned to desired wavelength ranges for excitationand/or emission. This may be useful, for example, in developing aparticular imaging technique using a known excitation source. By way ofexample, compounds of the formulas (FX1) to (FX18) may be modified toinclude one or more electron withdrawing substituents (EWG) on one ormore positions. Compounds of the formulas (FX1) to (FX18) may bemodified to include one or more electron donating substituents (EDG) onone or more positions. One class of electron withdrawing substituents(EWG) is: —CN, —CO₂R¹, —CONR²R³, —COR⁴, —NO₂, —SOR⁵, —SO₂R⁶, —SO₂OR⁷,—SO₂NR⁸R⁹, or —PO₃R¹⁰R¹¹. One class of electron donating substituents(EDG) is: —OR¹², —SR¹³, —NR¹⁴R¹⁵, —NR¹⁶COR¹⁷, and —P(R¹⁸) (where each ofR¹ to R¹⁸ are independently hydrogen or C₁-C₆ alkyl).

In an aspect of the invention, adding an EDG on an odd-numberedsubstituent of a formula of the invention will lead to a red shift inthe wavelength of light emitted from the compound. In an aspect of theinvention, adding an EWG on an odd-numbered substituent will lead to ablue shift in the wavelength of light emitted from the compound. In anaspect of the invention, adding an EDG on an even-numbered substituentwill lead to a blue shift in the wavelength of light emitted from thecompound. In an aspect of the invention, adding an EWG on aneven-numbered substituent will lead to a red shift in the wavelength oflight emitted from the compound. This information can be used to tailorthe spectral properties of the compounds of the invention.

Example 3. Phototherapy Using Phototherapeutic Analogs

Phototherapy, such as photodynamic therapy (PDT), typically employs acombination of a nontoxic photosensitizer (PS) and visible or nearinfrared light to generate reactive species that kill or otherwisedegrade target cells, such as tumors or other lesions. The presentinvention provides phototherapeutic agents useful for phototherapy.

Compounds of the invention having one or more photoreactive moieties orphotoreactive moiety generating groups (“phototherapeutic analogs” or“phototherapeutic agents”) are useful for phototherapeutic applications.Photoreactive moieties include, but are not limited to free radicals,carbenes, nitrenes, singlet oxygen, and the like. Examples of Type Iphotoreactive moieties that can be incorporated into a non-benzenoidaromatic compound for the purpose of synthesizing a phototherapeuticanalog include, but are not limited to, azides, azo compounds, diazocompounds, sulfenates, thiadiazoles, peroxides, and the free radical orreactive intermediate formed upon irradiation. Examples of Type IIphotoreactive moieties that can be incorporated into a non-benzenoidaromatic compound for the purpose of synthesizing a phototherapeuticanalog include, but are not limited to, phthalocyanines, porphyrins,extended porphyrins, and benzoporphyrins.

In aspects of this embodiment, compounds of the formulas (FX1) to (FX18)contain one or more functional groups that produce reactiveintermediates such as those photoreactive moieties listed above. Inaspects of this embodiment, compounds of the formulas (FX1) to (FX18)include one or more azides, azo compounds, diazo compounds, sulfenates,thiadiazoles, or peroxides.

In an embodiment, a separate Type 1 or Type 2 phototherapeutic agent isco-administered to the patient with a non-benzenoid aromatic compound ofthe invention. The methods described herein fully contemplate thisembodiment, in which case the term “phototherapeutic agent” isunderstood to include a non-benzenoid aromatic compound and a separateType 1 or Type 2 phototherapeutic agent, which can be administeredseparately or together.

In some embodiments, for example, the present invention provides methodscomprising administration of a phototherapeutic agent. By way ofexample, the phototherapeutic agent is administered to a patient.Administration provides delivery of the phototherapeutic agent to atarget tissue, such as a tumor. The patient may be optionally imaged asdescribed elsewhere to determine the location where the compound isbound within the patient. Once the compound is determined to be bound tothe targeted site or sites, the phototherapeutic agent is irradiatedwith a wavelength and intensity of light (for example electromagneticradiation having wavelengths selected over the range of 400 nm to 1300nm) sufficient to cause photofragmentation of the phototherapeuticagent. The photofragmentation typically results in homolytic cleavage ofthe agent, resulting in the generation of free radical intermediates.The generated free radicals then damage diseased tissues or cells of thetargeted site(s) to which the phototherapeutic agent had bound, therebytherapeutically treating the condition of the patient.

In an embodiment, the invention provides a method for a phototherapeuticprocedure, the method comprising: (i) administering to a patient aneffective amount of a phototherapeutic agent having the formula

wherein:

X_(a) is N or —C-(L⁶)_(q)-W⁶—R⁶;

X_(b) is N or —C-(L⁷)_(q)-W⁷—R⁷;

X_(c) is N or —C-(L⁸)_(q)-W⁸—R⁸;

X_(d) is N or —C-(L¹)_(q)-W¹—R¹;

X_(e) is N or —C-(L²)_(q)-W²—R²;

each of L¹ to L⁸ is independently C₁-C₁₀ alkylene, C₃-C₁₀ cycloalkylene,C₂-C₁₀ alkenylene, C₃-C₁₀ cycloalkenylene, C₂-C₁₀ alkynylene,ethenylene, ethynylene, phenylene, 1-aza-2,5-dioxocyclopentylene,—(CH₂CH₂O)_(m)—, —(CHOH)_(m)—, or 1,4-diazacyclohexylene;

each m is independently an integer selected from the range of 1 to 100;

each q is independently 0 or 1;

each of W¹ to W⁸ is independently a single bond, —(CH₂)_(n)—,—O(CH₂)_(n)—, —(CH₂)_(n)O—, —(HCCH)_(n)—, —O—, —S—, —SO—, —SO₂—, —SO₃ ⁻,—OSO₂—, —NR⁹—, —CO—, —COO—, —OCO—, —OCOO—, —CONR¹⁰—, —NR¹¹CO—,—OCONR¹²—, —NR¹³COO—, —NR¹⁴CONR¹⁵—, —NR¹⁶CSNR¹⁷—, —O(CH₂)_(n)—,—S(CH₂)_(n)—, —NR¹⁸(CH₂)_(n)—, —CO(CH₂)_(n)—, —COO(CH₂)_(n)—,—OCO(CH₂)_(n)—, —OCOO(CH₂)_(n)—, —CONR¹⁹(CH₂)_(n)—,—CONR²⁰(CH₂)_(n)(OCH₂CH₂)_(u)—, —NR²¹CO(CH₂)_(n)—, —OCONR²²(CH₂)_(n)—,—NR²³COO(CH₂)_(n)—, —NR²⁴CONR²⁵(CH₂)_(n)—, —NR²⁶CSNR²⁷(CH₂)_(n)—,—O(CH₂)_(n)NR²⁸CO(CH₂)_(n)—,—CO(CH₂)_(n)(CH₂OCH₂)_(n)(CH₂)_(n)NR²⁹(CH₂)_(n)NR³⁰CO—, —NR⁶⁹SR⁷⁰—, or—CO(CH₂)_(n)NR³¹CO—;

each n is independently selected from the range of 1 to 10;

each of R⁹ to R³¹ and each of R⁶⁹ to R⁷⁰ is independently hydrogen,C₁-C₂₀ alkyl, or C₅-C₃₀ aryl;

each of R¹ to R⁸ is independently hydrogen, C₁-C₂₀ alkyl, C₅-C₃₀ aryl,C₁-C₂₀ acyl, C₂-C₂₀ alkenyl, C₂-C₂₀ alkynyl, C₅-C₂₀ alkylaryl, C₁-C₆alkoxycarbonyl, halo, halomethyl, dihalomethyl, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷, —SO₂R³⁸, —SO₂OR³⁹,—SO₂NR⁴⁰R⁴¹, —PO₃R⁴²R⁴³, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, —NR⁴⁸COR⁴⁹,—CH₂(CHOH)_(n)R⁵⁰, —(CH₂CH₂O)_(u)R⁵¹, —CH(R⁵²)CO₂H, —CH(R⁵³)NH₂, TG¹ toTG⁸, PS¹ to PS⁸, or FL¹ to FL⁸;

each u is independently an integer selected from the range of 1 to 25;

each of R³² to R⁵⁵ is independently hydrogen or C₁-C₁₀ alkyl;

each of TG¹ to TG⁸ is independently an amino acid, a peptide, a protein,a nucleoside, a nucleotide, an enzyme, a carbohydrate, a glycomimetic,an oligomer, a lipid, a polymer, an antibody, an antibody fragment, amono- or polysaccharide comprising 1 to 50 carbohydrate units, aglycopeptide, a glycoprotein, a peptidomimetic, a drug, a drug mimic, ahormone, a receptor, a metal chelating agent, a radioactive ornonradioactive metal complex, a mono- or polynucleotide comprising 1 to50 nucleic acid units, a polypeptide comprising 2 to 30 amino acidunits, or an echogenic agent;

each of PS¹ to PS⁸ is independently a photosensitizing moiety capable ofproducing one or more free radicals, nitrenes, carbenes, and/or singletoxygen, and wherein each of PS¹ to PS⁸ comprises at least one azide,azo, diazo, oxaza, diaza, dithia, thioxa, dioxa, phthalocyanine,rhodamine, or porphyrin group; and

each of FL¹ to FL⁸ is independently a fluorescent group corresponding toa naphthoquinone, an anthracene, an anthraquinone, a phenanthrene, atetracene, a naphthacenedione, a pyridine, a quinoline, an isoquinoline,an indole, an isoindole, a pyrrole, an imidiazole, a pyrazole, apyrazine, a purine, a benzimidazole, a benzofuran, a dibenzofuran, acarbazole, an acridine, an acridone, a phenanthridine, a thiophene, abenzothiophene, a dibenzothiophene, a xanthene, a xanthone, a flavone, acoumarin, a phenoxazine, a phenothiazine, a phenoselenazine, a cyanine,an indocyanine, or an azo compound;

wherein any adjacent R¹ to R⁸ may combine with one or two optional—CR⁵⁴R⁵⁵— groups to form C₅-C₆ cycloalkyl, C₅-C₆ heterocycloalkyl, C₆aryl or C₅-C₆ heteroaryl; and (ii) exposing the phototherapeutic agentadministered to the patient to electromagnetic radiation. In anembodiment of this method, at least one of R¹ to R⁸ is C₁-C₁₀ alkyl,—OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹; and wherein at least one of R¹ toR⁸ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂,—SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₁-C₁₀ acyl; or wherein at least one of-(L¹)_(q)-W¹—R¹, -(L²)_(q)-W²—R², -(L³)_(q)-W³—R³, -(L⁴)_(q)-W⁴—R⁴,-(L⁵)_(q)-W⁵—R⁵, -(L⁶)_(q)-W⁶—R⁶, -(L⁷)_(q)-W⁷—R⁷, or -(L)_(q)-W⁸—R⁸includes a —(OCH₂CH₂)_(u)— group.

In an embodiment, the method of this aspect of the invention comprisesadministering to a patient a compound having any one of formulae (FX1)to (FX17), including all of the specific compositions classes andcompounds described in connection with formula (FX1) to (FX17), incombination with any of the methods steps presented herein. In anembodiment, the method of this aspect of the invention comprisesadministering to a patient a compound having formula (FX18), includingall of the specific compositions classes and compounds described inconnection with formula (FX18), in combination with any of the methodssteps presented herein.

In an embodiment, included is a non-benzenoid compound of any of theformulas described herein including formulae (FX1) to (FX17) wherein thecompound is a Type 2 photosensitizer without a photosensitizing groupattached. In an embodiment, included is a non-benzenoid compound of anyof the formulas described herein including formula (FX18) wherein thecompound is a Type 2 photosensitizer without a photosensitizing groupattached.

Phototherapy methods of the invention include photodynamic therapy andthermal laser photocoagulation. Methods of the present invention mayfurther comprise a number of additional steps. In an embodiment, forexample, the method further comprises delivering a phototherapeuticagent of any one of formulae (FX1) to (FX17) to a target tissue ororgan, such as a tumor. In an embodiment, the method further comprisescontacting a target tissue or organ with a phototherapeutic agent of anyone of formulae (FX1) to (FX17). In an embodiment, the present methodsfurther comprise the step of administering phototherapeutic agent of anyone of formulae (FX1) to (FX17) into a bodily fluid of the subject.Phototherapy methods of the invention include photodynamic therapy andthermal laser photocoagulation. Methods of the present invention mayfurther comprise a number of additional steps. In an embodiment, forexample, the method further comprises delivering a phototherapeuticagent of formula (FX18) to a target tissue or organ, such as a tumor. Inan embodiment, the method further comprises contacting a target tissueor organ with a phototherapeutic agent of formula (FX18). In anembodiment, the present methods further comprise the step ofadministering phototherapeutic agent of formula (FX18) into a bodilyfluid of the subject. Phototherapeutic agents may be introduced into thepatient by any suitable method, including intravenous, intraperitonealor subcutaneous injection or infusion, oral administration, transdermalabsorption through the skin, or by inhalation. In an embodiment, themethod further comprises allowing the phototherapeutic agent toaccumulate in a target tissue or organ prior to exposure of thephototherapeutic agent to electromagnetic radiation. In an embodiment,the phototherapeutic agent is administered to the skin, a tumor,surgical site, or a wound site. In an embodiment, for example, thephototherapeutic agent is administered and/or delivered to a bloodvessel, lung, heart, throat, ear, rectum, bladder, stomach, intestines,esophagus, liver, brain, prostrate, breast or pancreas of the subject.

In an embodiment, a therapeutically effective amount of thephototherapeutic agent is provided to the subject. For example,parenteral administration advantageously contains a sterile aqueoussolution or suspension of the phototherapeutic agent having aconcentration of an active agent comprising the compound of any one offormulae (FX1) to (FX18) ranging from about 0.0 μM to about 0.5M.Preferred parenteral formulations have a concentration of an activeagent comprising the compound of any one of formulae (FX1) to (FX18)selected over the range of 1 μM to 10 mM. Such solutions also maycontain pharmaceutically acceptable buffers, emulsifiers, surfactants,and, optionally, electrolytes such as sodium chloride. The dose of thecompound of any one of formulae (FX1) to (FX18) may vary from 0.1 to 500mg/kg body weight, preferably from 0.5 to 2 mg/kg body weight.

In methods of the present invention, the phototherapeutic agent and anyother components can be formulated for enteral (oral or rectal),parenteral, topical, or cutaneous administration. Topical or cutaneousdelivery of the phototherapeutic agent may also include aerosols,creams, gels, solutions, emulsions and colloids. The compositions areadministered in doses effective to achieve the desired diagnostic ortherapeutic objective. Such doses may vary widely depending upon theparticular complex employed, the organs or tissues to be examined ortreated, the equipment employed in the clinical procedure, the efficacyof the treatment achieved, and the like. These compositions contain aneffective amount of the phototherapeutic agent along with conventionalpharmaceutical carriers and excipients appropriate for the type ofadministration contemplated. These compositions may also includestabilizing agents and skin penetration enhancing agents and also maycontain pharmaceutically acceptable buffers, emulsifiers, surfactants,and, optionally, electrolytes such as sodium chloride. Formulations forenteral administration may vary widely, as is well known in the art. Ingeneral, such formulations are liquids, which include an effectiveamount of the complexes in aqueous solution or suspension. Such enteralcompositions may optionally include buffers, surfactants, emulsifiers,thixotropic agents, and the like. Compounds for oral administration mayalso contain flavoring agents and other ingredients for enhancing theirorganoleptic qualities. Formulations for topical delivery may alsocontain liquid or semisolid excipients to assist in the penetration ofthe photosensitizer. The compounds may also be delivered in an aerosolspray.

As will be understood by one having skill in the art, the opticalconditions for the step of exposing the phototherapeutic agentadministered to the patient to electromagnetic radiation will varyconsiderably with the (i) therapeutic and/or diagnostic objectives, and(ii) the condition of the subject (e.g., height, weight, state of healthetc.). In an embodiment, electromagnetic radiation has wavelengths,energy and/or fluence sufficient to achieve a desired therapeutic and/ordiagnostic result. In an embodiment, the electromagnetic radiation haswavelengths, energy and/or fluence sufficient to activate thephototherapeutic agent. In a method, the electromagnetic radiationexposed to the phototherapeutic agent has wavelengths selected over therange of 400 nm-1300 nm, preferably for some applications 400 nm-900 nm,preferably for some applications 500 nm to 900 nm. In an embodiment, theelectromagnetic radiation exposed to the phototherapeutic agent haswavelengths corresponding to a maximum in the absorption spectrum of thephototherapeutic agent, preferably for some applications a maximum inthe visible or NIR regions of the electromagnetic spectrum. Optionally,excitation is achieved using electromagnetic substantially free (e.g.,less than about 10% of total radiant energy), of ultraviolet radiation,for example, to minimize exposure of the subject to electromagneticradiation capable of causing unwanted cell or tissue damage. As will beappreciated by one having skill in the art, the fluence employed duringexcitation of the phototherapeutic agent can vary depending on the typeof tissue, depth of the target, composition of the phototherapeuticagent and the amount on composition of overlying fluid and blood. Insome embodiments, the fluence employed is selected over the range of 10to 500 Joules cm⁻². The irradiance is typically selected from the rangeof 50 to 1500 mWcm⁻², preferably of 50 to 500 mWcm⁻². Electromagneticradiation may be provided to the phototherapeutic agent using a range ofoptical sources and/or surgical instrumentation, including a laser,light emitting diodes, fiber optic device, endoscope, catheter, opticalfilters, or any combination of these.

Example 4: Biotargeting Using Non-Benzenoid Aromatic Compounds

Compounds of the invention are also useful for targeting biologicalmoieties. Targeted moieties may also undergo subsequent or coincidentphototherapeutic applications.

In aspects of this embodiment, compounds of the formulas (FX1) to (FX18)contain one or more biotargeting groups. These ligands are well known inthe art. By way of example, the non-benzenoid compound which includes atargeting moiety can be administered to a patient in a diagnosticallyeffective amount to detect the non-benzenoid compound within thepatient. After a period of time has lapsed for the compound to bind tothe desired target, the whole body or portion thereof is exposed tolight of suitable wavelength to excite the non-benzenoid compound. Lightemanating from the patient as a result of the absorption and excitationof the non-benzenoid compound is then detected. By evaluating thelocation and strength of light emanating from the patient, a diagnosiscan be made as a result of the targeting properties of the non-benzenoidcompound.

In embodiments, compounds of the invention are useful for both oncologyand non-oncology applications. Some specific targets are tumorsaccessible via endoscope. In this application, a compound that targets apeptide associated with such a tumor is administered to the tumor viaendoscope or other useful method. Then, the compounds of the inventioncan be used in phototherapeutic applications or imaging applications.Other specific targets include colon, lung, ovarian, cervical,esophageal, bladder, blood, and stomach cancers; endometriosis, andbacterial infections. Particular targeting groups include ST receptorbinding agents, bombesin receptor binding agents, leukemia peptides, andfolate receptor binding. Some examples of targeting peptides aredescribed in WO/2008/108941.

In one example, a targeted compound can contain all or part of a steroidhormone or a steroid receptor binding compound, and therefore targetsteroid hormone sensitive receptors. In this example, the targetedcompound is administered, targets and preferably accumulates in thedesired site such as breast and/or prostate lesion and is photoactivatedfor monitoring, imaging, or therapy remotely or at the target site.Similar target binding molecules and uses will be recognized by oneskilled in the art. For example, the targeted compound can be a compoundthat targets and binds to a somatostatin, bombesin, CCK, and/orneurotensin receptor binding molecule, or can be a carcinogenicembryonic antigen-binding compound that binds to a carcinogenicembryonic antigen. These are then photoactivated at, for example, lungcancer cells with CCK receptor binding molecules, colorectal cancercells with ST receptor and carcinoembryonic antigen (CEA) bindingmolecules, melanoma cells with dihydroxyindolecarboxylic acid, vascularsites of atherosclerotic plaque with integrin receptor bindingmolecules, brain lesions with amyloid plaque binding molecules, and thelike.

Successful specific targeting of photoactive compounds to tumors usingantibodies and peptides for diagnostic imaging of tumors has beendescribed in Achilefu et al., Novel receptor-targeted fluorescentcontrast agents for in vivo imaging of tumors, Investigative Radiology,2000, 35, pp. 479-485; Ballou et al., Tumor labeling in vivo usingcyanine conjugated monoclonal antibodies, Cancer Immunology andImmunotherapy, 1995, 41, pp. 257-263; and Licha et al., New contrastagent for optical imaging: acid cleavable conjugates of cyanine dyeswith biomolecules, in Biomedical Imaging: Reporters, Dyes andInstrumentation, Proceedings of SPIE, 1999, 3600, pp. 29-35. As such, itis widely accepted that targeted photochemicals are effective intargeting, detecting and treating a wide range of physiological andbiological sites.

The optical agents of this example can contain additionalfunctionalities that can be used to attach various types ofbiomolecules, synthetic polymers, and organized aggregates for selectivedelivery to various organs or tissues of interest. Examples of syntheticpolymers include polyaminoacids, polyols, polyamines, polyacids,oligonucleotides, aborols, dendrimers, and aptamers. The inventionincludes, but is not limited to, phototherapeutic agents comprising anoptical agent—biomolecule conjugate which provides advantages overnonspecific optical agents or the conjugation of optical agents to verylarge biomolecules. These conjugates provide enhanced localization in,and rapid visualization of, tumors which is beneficial for imaging,monitoring, diagnosis and therapy. The agents are rapidly cleared fromblood and non-target tissues so there is less concern for accumulationand for toxicity. A variety of high purity compounds can be easilysynthesized for combinatorial screening of new targets, e.g., toidentify receptors or targeting agents, and for the ability to affectthe pharmacokinetics of the conjugates by minor structural changes.

In some embodiments, a liposome or micelle can be utilized as a carrieror vehicle for the composition. For example, in some embodiments, anoptical agent comprises a compound of the invention that can be a partof the lipophilic bilayers or micelle, and the targeting ligand, ifpresent, can be on the external surface of the liposome or micelle. Asanother example, a targeting ligand can be externally attached to theliposome or micelle after formulation for targeting the liposome ormicelle (which contains a phototherapeutic agent/photosensitizercompound of the invention) to the desired tissue, organ, or other sitein the body.

In embodiments, compounds of the invention are useful for both oncologyand non-oncology applications. Some specific targets are tumorsaccessible via endoscope. In an application, a compound that targets aprotein, polypeptide, oligonucleotide or other biomolecule associatedwith such a tumor is administered to the tumor via endoscope or otheruseful method. Then, the compounds of the invention can be used inphototherapeutic applications, monitoring applications, diagnosisapplications or imaging applications. Other specific targets includecolon, lung, ovarian, cervical, esophageal, bladder, blood, stomachcancers, endometriosis, and bacterial infections.

Targeting Ligands

The estrogen receptor is an example of a steroid receptor to whichsteroid receptor binding molecules would bind. The following compoundsare known to bind to the estrogen receptor: estratriol;17β-aminoestrogen (AE) derivatives such as prolame and butolame; drugssuch as tamoxifen, ICI-164384, raloxifene, and genistein; 17β-estradiol;glucocorticoids; progesterone; estrogens; retinoids; fatty acidderivatives; and phytoestrogens. In addition, commercially availablekits can identify compounds specific for binding to the estrogenreceptor (e.g., Estrogen Receptor-alpha Competitor Assay Kit, Red; andEstrogen Receptor-beta Competitor Assay Kit, Red (Invitrogen Corp.,Carlsbad Calif.).

The glucose receptor is an example of a carbohydrate receptor to whichcarbohydrate receptor binding molecules would bind. The glucoseconjugate N-palmitoyl glucosamine [NPG] is known to bind the glucosereceptor (Dufes et al., Pharm. Res. 17:1250, 2000). The glycoproteinhormone receptor is another example of a carbohydrate receptor to whichcarbohydrate receptor binding molecules would bind. Follicle stimulatinghormone (FSH) is known to bind the glycoprotein hormone receptor (Tillyet al., Endocrinology 131: 799, 1992). Other compounds known to bind thecarbohydrate receptor, and hence examples of carbohydrate receptorbinding molecules, are: polysialic acid, bacterial adhesins (specializedsurface proteins that mediate binding of many pathogenic bacteria, suchas enterohemorrhagic E. coli (EHEC) and Shigella dysenteriae, to hostcells, which allow these bacteria to colonize host cell surfaces),soluble carbohydrate receptor analogs, artificial glycopolymers andother multivalent glycoconjugates such as an acrylamide copolymercarrying -L-fucopyranoside and 3-sulfo-D-galactopyranoside in clusters,isomeric carbohydrates, synthetic derivatives, neoglycoproteins,neoglycolipids, glycosidases, and glycosyltransferases. Carbohydratebinding proteins can be screened with phage display libraries as knownto a person of ordinary skill in the art.

Somatostatin receptor binding molecules include somatostatin andsomatostatin receptor analogs, octreotide, glycosylated somatostatin-14(somatostatin-dextran⁷⁰), seglitide, and peptides P587 and P829 asdescribed in Vallabhajosula et al., J. Nuclear Med., 37:1016, 1996.

Cholecystokinin receptor binding molecules include the endogenouspeptides cholecystekinin (CCK)-4, CCK-8, CCK-33, and gastrin;antagonists devazepide and lorglumide; agonists BC264[Tyr(SO₃H)-gNle-mGly-Trp-(NMe)Nle-Asp-Phe-NH₃] and desulfated CCK-8;Kinevac (synthetic cholecystekinin, sincalide); and CCK analoguesmodified at the sulfated tyrosyl at position 27.

Neurotensin receptor binding molecules include neurotensin, neuromedinN, JMV449 (H-Lysψ(CH₂NH)-Lys-Pro-Tyr-Ile-Leu), the non-peptideantagonist SR142948A(2-([5-(2,6-dimethoxyphenyl)-1-(4-(N-[3-dimethylaminopropyl]-N-methyl]-N-methylcarbamoyl)-2-isopropylphenyl)-1H-pyrazole-3-carbonyl)amino)adamantine-2-carboxylicacid hydrochloride), and levocobastine. Commercially availableneurotensin receptor binding kits can evaluate potential neurotensinreceptor binding molecules (e.g., DELFIA Neurotensin Receptor BindingKit, PerkinElmer (Boston Mass.)).

Bombesin receptor binding molecules include the endogenous ligandsgastrin-releasing peptide (GRP), neuromedin B (NMB), and GRP-18-27, andantagonists including JMV-1458 (glycine-extended bombesin(paraphydroxy-phenyl-propionyl-Gln-Trp-Ala-Val-Gly-His-Leu-Met-Gly-OH)),JMV-641, JMV-1799, and JMV-1802, PD165929,1-naphthoyl-[DAla²⁴,DPro²⁶,ψ26-27]GRP-20-27, kuwanon H, and kuwanon G.Commercially available bombesin receptor binding kits can evaluatepotential bombesin receptor binding molecules (e.g., DELFIA BombesinReceptor Binding Kit, PerkinElmer (Boston Mass.)).

ST receptor binding molecules include native ST peptide, and SEQ IDNO:2, SEQ ID NO:3, SEQ ID NOS:5-54 and fragments and derivatives thereoffrom U.S. Pat. No. 5,518,888.

Compounds of the invention can contain all or part of a targetingligand, receptor or peptide known to bind to a specific target. Methodsof attaching the targeting ligand, receptor or peptide to a compound ofthe invention are described elsewhere herein.

Example 5: Administration and Formulation

Salts and Prodrugs

The invention contemplates pharmaceutically active compounds eitherchemically synthesized or formed by in vivo biotransformation tocompounds set forth herein.

Optical agents of the invention can be formulated withpharmaceutically-acceptable anions and/or cations. Pharmaceuticallyacceptable salts comprise pharmaceutically-acceptable anions and/orcations. As used herein, the term “pharmaceutically acceptable salt” canrefer to acid addition salts or base addition salts of the compounds inthe present disclosure. A pharmaceutically acceptable salt is any saltwhich retains at least a portion of the activity of the parent compoundand does not impart significant deleterious or undesirable effect on asubject to whom it is administered and in the context in which it isadministered. Pharmaceutically acceptable salts include metal complexesand salts of both inorganic and organic acids. Pharmaceuticallyacceptable salts include metal salts such as aluminum, calcium, iron,magnesium, manganese and complex salts. Pharmaceutically acceptablesalts include, but are not limited to, acid salts such as acetic,aspartic, alkylsulfonic, arylsulfonic, axetil, benzenesulfonic, benzoic,bicarbonic, bisulfuric, bitartaric, butyric, calcium edetate, camsylic,carbonic, chlorobenzoic, cilexetil, citric, edetic, edisylic, estolic,esyl, esylic, formic, fumaric, gluceptic, gluconic, glutamic, glycolic,glycolylarsanilic, hexamic, hexylresorcjnoic, hydrabamic, hydrobromic,hydrochloric, hydroiodic, hydroxynaphthoic, isethionic, lactic,lactobionic, maleic, malic, malonic, mandelic, methanesulfonic,methylnitric, methylsulfuric, mucic, muconic, napsylic, nitric, oxalic,p-nitromethanesulfonic, pamoic, pantothenic, phosphoric, monohydrogenphosphoric, dihydrogen phosphoric, phthalic, polygalactouronic,propionic, salicylic, stearic, succinic, sulfamic, sulfanlic, sulfonic,sulfuric, tannic, tartaric, teoclic, toluenesulfonic, and the like.Pharmaceutically acceptable salts can be derived from amino acids,including, but not limited to, cysteine. Other pharmaceuticallyacceptable salts can be found, for example, in Stahl et al., Handbook ofPharmaceutical Salts: Properties, Selection, and Use, Wiley-VCH, VerlagHelvetica Chimica Acta, Zürich, 2002. (ISBN 3-906390-26-8).

Efficacy

Toxicity and therapeutic efficacy of such compounds and bioconjugatescan be determined by standard pharmaceutical procedures in cell culturesor experimental animals for determining the LD₅₀ (the dose lethal to 50%of the population) and the ED₅₀, (the dose therapeutically effective in50% of the population). The dose ratio between toxic and therapeuticeffects is the therapeutic index that can be expressed as the ratioLD₅₀/ED₅₀. Compounds and bioconjugates that exhibit large therapeuticindices are preferred. While compounds and bioconjugates exhibitingtoxic side effects can be used, care should be taken to design adelivery system that targets such compounds and bioconjugates to thesite affected by the disease or disorder in order to minimize potentialdamage to unaffected cells and reduce side effects.

Data obtained from the cell culture assays and animal studies can beused in formulating a range of dosages for use in humans and othermammals. The dosage of such compounds and bioconjugates lies preferablywithin a range of circulating plasma or other bodily fluidconcentrations that include the ED₅₀ and provides clinically efficaciousresults (i.e., reduction in disease symptoms). The dosage can varywithin this range depending upon the dosage form employed and the routeof administration utilized. For any compound and bioconjugate of thepresent invention, the therapeutically effective amount can be estimatedinitially from cell culture assays. A dosage can be formulated in animalmodels to achieve a circulating plasma concentration range that includesthe ED₅₀ (the concentration of the test compound that achieves ahalf-maximal inhibition of symptoms) as determined in cell culture. Suchinformation can be used to more accurately determine useful dosages inhumans and other mammals. Compound and bioconjugate levels in plasma canbe measured, for example, by high performance liquid chromatography.

An amount of a compound or bioconjugate that can be combined with apharmaceutically acceptable carrier to produce a single dosage form willvary depending upon the patient treated and the particular mode ofadministration. It will be appreciated by those skilled in the art thatthe unit content of a compound/bioconjugate contained in an individualdose of each dosage form need not in itself constitute a therapeuticallyeffective amount, as the necessary therapeutically effective amountcould be reached by administration of a number of individual doses. Theselection of dosage depends upon the dosage form utilized, the conditionbeing treated, and the particular purpose to be achieved according tothe determination of those skilled in the art.

The dosage and dosage regime for treating a disease or condition can beselected in accordance with a variety of factors, including the type,age, weight, sex, diet and/or medical condition of the patient, theroute of administration, pharmacological considerations such asactivity, efficacy, pharmacokinetic and/or toxicology profiles of theparticular compound/bioconjugate employed, whether acompound/bioconjugate delivery system is utilized, and/or whether thecompound/bioconjugate is administered as a pro-drug or part of a drugcombination. Thus, the dosage regime actually employed can vary widelyfrom subject to subject, or disease to disease and different routes ofadministration can be employed in different clinical settings.

The identified compounds/bioconjugates monitor, treat, inhibit, controland/or prevent, or at least partially arrest or partially prevent,diseases and conditions of interest and can be administered to a subjectat therapeutically effective amounts and optionally diagnosticallyeffective amounts. Compositions/formulations of the present inventioncomprise a therapeutically effective amount (which can optionallyinclude a diagnostically effective amount) of at least one compound orbioconjugate of the present invention. Subjects receiving treatment thatincludes a compound/bioconjugate of the invention are preferably animals(e.g., mammals, reptiles and/or avians), more preferably humans, horses,cows, dogs, cats, sheep, pigs, and/or chickens, and most preferablyhumans.

Administration

The diagnostic and therapeutic formulations of this invention can beadministered alone, but can be administered with a pharmaceuticalcarrier selected upon the basis of the chosen route of administrationand standard pharmaceutical practice.

Any suitable form of administration can be employed in connection withthe diagnostic and therapeutic formulations of the invention. Thediagnostic and therapeutic formulations of this invention can beadministered intravenously, in oral dosage forms, intraperitoneally,subcutaneously, or intramuscularly, all using dosage forms well known tothose of ordinary skill in the pharmaceutical arts.

The present compositions, preparations and formulations can beformulated into diagnostic or therapeutic compositions for enteral,parenteral, topical, aerosol, inhalation, or cutaneous administration.Topical or cutaneous delivery of the compositions, preparations andformulations can also include aerosol formulation, creams, gels,solutions, etc. The present compositions, preparations and formulationsare administered in doses effective to achieve the desired diagnosticand/or therapeutic effect. Such doses can vary widely depending upon theparticular compositions employed in the composition, the organs ortissues to be examined, the equipment employed in the clinicalprocedure, the efficacy of the treatment achieved, and the like. Thesecompositions, preparations and formulations contain an effective amountof the composition(s), along with conventional pharmaceutical carriersand excipients appropriate for the type of administration contemplated.These compositions, preparations and formulations can also optionallyinclude stabilizing agents and skin penetration enhancing agents.

(i) Parenteral Administration

Compounds and bioconjugates of the present invention can be formulatedfor parenteral administration by injection (e.g., by bolus injection orcontinuous infusion). Formulations for injection can be presented inunit dosage form in ampoules or in multi-dose containers with anoptional preservative added. The parenteral preparation can be enclosedin ampoules, disposable syringes or multiple dose vials made of glass,plastic or the like. The formulation can take such forms as suspensions,solutions or emulsions in oily or aqueous vehicles, and can containformulatory agents such as suspending, stabilizing and/or dispersingagents.

For example, a parenteral preparation can be a sterile injectablesolution or suspension in a nontoxic parenterally acceptable diluent orsolvent (e.g., as a solution in 1,3-butanediol).

Among the acceptable vehicles and solvents that can be employed arewater, Ringer's solution, and isotonic sodium chloride solution. Inaddition, sterile, fixed oils are conventionally employed as a solventor suspending medium. For this purpose any bland fixed oil can beemployed including synthetic mono- or di-glycerides. In addition, fattyacids such as oleic acid can be used in the parenteral preparation.

Alternatively, compounds and bioconjugates of the present invention canbe formulated in powder form for constitution with a suitable vehicle,such as sterile pyrogen-free water, before use. For example, acompound/bioconjugate suitable for parenteral administration can includea sterile isotonic saline solution containing between 0.1 percent and 90percent weight per volume of the compound/bioconjugate. By way ofexample, a solution can contain from about 5 percent to about 20percent, more preferably from about 5 percent to about 17 percent, morepreferably from about 8 to about 14 percent, and still more preferablyabout 10 percent weight per volume of the compound/bioconjugate. Thesolution or powder preparation can also include a solubilizing agent anda local anesthetic such as lignocaine to ease pain at the site of theinjection. Other methods of parenteral delivery ofcompounds/bioconjugates will be known to the skilled artisan and arewithin the scope of the invention.

(ii) Oral Administration

For oral administration, a compound/bioconjugate of the invention can beformulated to take the form of tablets or capsules prepared byconventional means with one or more pharmaceutically acceptable carriers(e.g., excipients such as binding agents, fillers, lubricants anddisintegrants).

(iii) Controlled-Release Administration

Controlled-release (or sustained-release) preparations can be formulatedto extend the activity of a compound/bioconjugate and reduce dosagefrequency. Controlled-release preparations can also be used to effectthe time of onset of action or other characteristics, such as bloodlevels of the compound/bioconjugate, and consequently affect theoccurrence of side effects.

Controlled-release preparations can be designed to initially release anamount of a compound/bioconjugate that produces the desired therapeuticeffect, and gradually and continually release other amounts of thecompound/bioconjugate to maintain the level of therapeutic effect overan extended period of time. In order to maintain a near-constant levelof a compound/bioconjugate in the body, the compound/bioconjugate can bereleased from the dosage form at a rate that will replace the amount ofcompound/bioconjugate being metabolized and/or excreted from the body.The controlled-release of a compound/bioconjugate can be stimulated byvarious inducers, e.g., change in pH, change in temperature, enzymes,water, and/or other physiological conditions or molecules.

Controlled-release systems can include, for example, an infusion pumpwhich can be used to administer the compound/bioconjugate in a mannersimilar to that used for delivering insulin or chemotherapy to the bodygenerally, or to specific organs or tumors. Typically, using such asystem, the compound/bioconjugate is administered in combination with abiodegradable, biocompatible polymeric implant that releases thecompound/bioconjugate over a controlled period of time at a selectedsite. Examples of polymeric materials include polyanhydrides,polyorthoesters, polyglycolic acid, polylactic acid, polyethylene vinylacetate, and copolymers and combinations thereof. In addition, acontrolled release system can be placed in proximity of a therapeutictarget (e.g., organ, tissue, or group of cells), thus requiring only afraction of a systemic dosage.

Compounds/bioconjugates of the invention can be administered by othercontrolled-release means or delivery devices that are well known tothose of ordinary skill in the art. These include, for example,hydropropylmethyl cellulose, other polymer matrices, gels, permeablemembranes, osmotic systems, multilayer coatings, microparticles,liposomes, microspheres, or the like, or a combination of any of theabove to provide the desired release profile in varying proportions.Other methods of controlled-release delivery of compounds/bioconjugateswill be known to the skilled artisan and are within the scope of theinvention.

(iv) Inhalation Administration

Compounds/bioconjugates of the invention can be administered directly tothe lung of a patient/subject by inhalation. For administration byinhalation, a compound/bioconjugate can be conveniently delivered to thelung by a number of different devices. For example, a Metered DoseInhaler (“MDI”) which utilizes canisters that contain a suitable lowboiling point propellant, e.g., dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide orother suitable gas can be used to deliver a compound/bioconjugatedirectly to the lung. MDI devices are available from a number ofsuppliers such as 3M Corporation, Aventis, Boehringer Ingleheim, ForestLaboratories, GlaxoSmithKline, Merck & Co. and Vectura.

Alternatively, a Dry Powder Inhaler (DPI) device can be used toadminister a compound/bioconjugate to the lung. DPI devices typicallyuse a mechanism such as a burst of gas to create a cloud of dry powderinside a container, which can then be inhaled by the patient. DPIdevices are also well known in the art and can be purchased from anumber of vendors which include, for example, GlaxoSmithKline, NektarTherapeutics, Innovata and Vectura. A popular variation is the multipledose DPI (“MDDPI”) system, which allows for the delivery of more thanone therapeutic dose. MDDPI devices are available from companies such asAstraZeneca, GlaxoSmithKline, TEVA, Merck & Co., SkyePharma and Vectura.For example, capsules and cartridges of gelatin for use in an inhaler orinsufflator can be formulated containing a powder mix of thecompound/bioconjugate and a suitable powder base such as lactose orstarch for these systems.

Another type of device that can be used to deliver acompound/bioconjugate to the lung is a liquid spray device supplied, forexample, by Aradigm Corporation. Liquid spray systems use extremelysmall nozzle holes to aerosolize liquid compound/bioconjugateformulations that can then be directly inhaled into the lung. Forexample, a nebulizer device can be used to deliver acompound/bioconjugate to the lung. Nebulizers create aerosols fromliquid compound/bioconjugate formulations by using, for example,ultrasonic energy to form fine particles that can be readily inhaled.Examples of nebulizers include devices supplied by Aventis and Battelle.

In another example, an electrohydrodynamic (“EHD”) aerosol device can beused to deliver a compound/bioconjugate to the lung. EHD aerosol devicesuse electrical energy to aerosolize liquid compound/bioconjugatesolutions or suspensions. The electrochemical properties of thecompound/bioconjugate formulation are important parameters to optimizewhen delivering this compound/bioconjugate to the lung with an EHDaerosol device. Such optimization is routinely performed by one of skillin the art. Other methods of intra-pulmonary delivery ofcompounds/bioconjugates will be known to the skilled artisan and arewithin the scope of the invention.

Liquid compound/bioconjugate formulations suitable for use withnebulizers and liquid spray devices and EHD aerosol devices willtypically include the compound/bioconjugate with a pharmaceuticallyacceptable carrier. In one exemplary embodiment, the pharmaceuticallyacceptable carrier is a liquid such as alcohol, water, polyethyleneglycol or a perfluorocarbon. Optionally, another material can be addedto alter the aerosol properties of the solution or suspension of thecompound/bioconjugate. For example, this material can be a liquid suchas an alcohol, glycol, polyglycol or a fatty acid. Other methods offormulating liquid compound/bioconjugate solutions or suspensionssuitable for use in aerosol devices are known to those of skill in theart.

(v) Depot Administration

A compound/bioconjugate of the invention can be formulated as a depotpreparation.

Such long-acting formulations can be administered by implantation (e.g.,subcutaneously or intramuscularly) or by intramuscular injection.Accordingly, the compound/bioconjugate can be formulated with suitablepolymeric or hydrophobic materials such as an emulsion in an acceptableoil or ion exchange resin, or as sparingly soluble derivatives such as asparingly soluble salt. Other methods of depot delivery ofcompounds/bioconjugates will be known to the skilled artisan and arewithin the scope of the invention.

(vi) Topical Administration

For topical application, a compound/bioconjugate can be combined with apharmaceutically acceptable carrier so that an effective dosage isdelivered, based on the desired activity ranging from an effectivedosage, for example, of 1.0 μM to 1.0 mM. In one aspect of theinvention, a topical formulation of a compound/bioconjugate can beapplied to the skin. The pharmaceutically acceptable carrier can be inthe form of, for example, and not by way of limitation, an ointment,cream, gel, paste, foam, aerosol, suppository, pad or gelled stick.

A topical formulation can include a therapeutically effective amount ofa compound/bioconjugate in an ophthalmologically acceptable excipientsuch as buffered saline, mineral oil, vegetable oils such as corn orarachis oil, petroleum jelly, Miglyol 182, alcohol solutions, orliposomes or liposome-like products. Any of these formulations of suchcompounds/bioconjugates can include preservatives, antioxidants,antibiotics, immunosuppressants, and other biologically orpharmaceutically effective agents that do not exert a significantdetrimental effect on the compound/bioconjugate. Other methods oftopical delivery of compounds/bioconjugates will be known to the skilledartisan and are within the scope of the invention.

(vii) Rectal Administration

Compounds/bioconjugates of the invention can be formulated in rectalformulations such as suppositories or retention enemas that includeconventional suppository bases such as cocoa butter or other glyceridesand/or binders and/or carriers such as triglycerides, microcrystallinecellulose, gum tragacanth or gelatin. Rectal formulations can contain acompound/bioconjugate in the range of 0.5% to 10% by weight, forexample. Other methods of rectal delivery of compounds/bioconjugateswill be known to the skilled artisan and are within the scope of theinvention.

(viii) Other Systems of Administration

Various other delivery systems are known in the art and can be used toadminister the compounds/bioconjugates of the invention. Moreover, theseand other delivery systems can be combined and/or modified to promoteoptimization of the administration of compounds/bioconjugates of thepresent invention. Exemplary formulations that includecompounds/bioconjugates of the present invention are described elsewhereherein (the compounds/bioconjugates of the present invention areindicated as the active ingredient, but those of skill in the art willrecognize that pro-drugs and compound combinations are also meant to beencompassed by this term).

Formulation

In an embodiment, the invention provides the use of one or morecompositions set forth herein for the diagnosis of a disease.

In an embodiment, the invention provides a pharmaceutical formulationhaving an active ingredient comprising a composition of the invention,such as a compound of any one of formulas (FX1)-(FX17). In anembodiment, the invention provides a method of synthesizing acomposition of the invention or a pharmaceutical formulation thereof,such as a compound of any one of formulas (FX1)-(FX17). In anembodiment, the invention provides a pharmaceutical formulation havingan active ingredient comprising a composition of the invention, such asa compound of any one of formulas (FX18). In an embodiment, theinvention provides a method of synthesizing a composition of theinvention or a pharmaceutical formulation thereof, such as a compound ofany one of formulas (FX18).

This invention also is directed, in part, to pharmaceutical compositionsincluding a therapeutically effective amount of a compound or salt ofthis invention, as well as processes for making such compositions. Suchcompositions generally include one or more pharmaceutically acceptablecarriers (e.g., excipients, vehicles, auxiliaries, adjuvants, diluents)and can include other active ingredients. Formulation of thesecompositions can be achieved by various methods known in the art. Ageneral discussion of these methods can be found in, for example,Hoover, John E., Remington's Pharmaceutical Sciences (Mack PublishingCo., Easton, Pa.: 1975). See also, Lachman, L., eds., PharmaceuticalDosage Forms (Marcel Decker, New York, N.Y., 1980).

The diagnostic and therapeutic formulations of this invention andmedicaments of this invention can further comprise one or morepharmaceutically acceptable carriers, excipients, buffers, emulsifiers,surfactants, electrolytes or diluents. Such compositions and medicamentsare prepared in accordance with acceptable pharmaceutical procedures,such as, for example, those described in Remingtons PharmaceuticalSciences, 17th edition, ed. Alfonoso R. Gennaro, Mack PublishingCompany, Easton, Pa. (1985).

Compositions of the invention include formulations and preparationscomprising one or more of the present compounds provided in an aqueoussolution, such as a pharmaceutically acceptable formulation orpreparation. Optionally, compositions of the invention further compriseone or more pharmaceutically acceptable surfactants, buffers,electrolytes, salts, carriers, binders, coatings, preservatives and/orexcipients.

Compounds and bioconjugates of the present invention can be formulatedby known methods for administration to a subject using several routeswhich include, but are not limited to, parenteral, oral, topical,intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous,intranasal, epidural, and ophthalmic routes. An individualcompound/bioconjugate can be administered in combination with one ormore additional compounds/bioconjugates of the present invention and/ortogether with other biologically active or biologically inert agents.Such biologically active or inert agents can be in fluid or mechanicalcommunication with the compound(s)/bioconjugate(s) or attached to thecompound(s)/bioconjugate(s) by ionic, covalent, Van der Waals,hydrophobic, hydrophilic or other physical forces. It is preferred thatadministration is localized in a subject, but administration can also besystemic.

Compounds and bioconjugates of the present invention can be formulatedby any conventional manner using one or more pharmaceutically acceptablecarriers. Thus, the compound(s)/bioconjugate(s) and theirpharmaceutically acceptable salts and solvates can be specificallyformulated for administration, e.g., by inhalation or insufflation(either through the mouth or the nose) or oral, buccal, parenteral orrectal administration. The compounds/bioconjugates can take the form ofcharged, neutral and/or other pharmaceutically acceptable salt forms.Examples of pharmaceutically acceptable carriers include, but are notlimited to, those described in REMINGTON'S PHARMACEUTICAL SCIENCES (A.R. Gennaro, Ed.), 20th edition, Williams & Wilkins PA, USA (2000).

Compounds and bioconjugates of the present invention can be formulatedin the form of solutions, suspensions, emulsions, tablets, pills,capsules, powders, controlled- or sustained-release formulations and thelike. Such formulations will contain a therapeutically effective amountof the compound/bioconjugate, preferably in purified form, together witha suitable amount of carrier so as to provide the form for properadministration to the patient. The formulation should suit the mode ofadministration.

In some embodiments, a liposome or micelle can be utilized as a carrieror vehicle for the composition. For example, in some embodiments, thecompound can be a part of the lipophilic bilayers or micelle, and thetargeting ligand, if present, can be on the external surface of theliposome or micelle. As another example, a targeting ligand can beexternally attached to the liposome or micelle after formulation fortargeting the liposome or micelle (which contains the optical agents) tothe desired tissue, organ, or other site in the body.

In one embodiment, the present compounds are formulated as nanoparticlesor microparticles. Use of such nanoparticle or microparticleformulations can be beneficial for some applications to enhancedelivery, localization, target specificity, administration, etc. of thecompound. Potentially useful nanoparticles and microparticles include,but are not limited to, micelles, liposomes, microemulsions,nanoemulsions, vesicles, tubular micelles, cylindrical micelles,bilayers, folded sheets structures, globular aggregates, swollenmicelles, inclusion complex, encapsulated droplets, microcapsules,nanocapsules or the like. As will be understood by those having skill inthe art, the present compounds can be located inside the nanoparticle ormicroparticle, within a membrane or wall of the nanoparticle ormicroparticle, or outside of (but bonded to or otherwise associatedwith) the nanoparticle or microparticle. The agent formulated innanoparticles or microparticles can be administered by any of the routespreviously described. In a formulation applied topically, the compoundis slowly released over time. In an injectable formulation, theliposome, micelle, capsule, etc., circulates in the bloodstream and isdelivered to the desired site (e.g., target tissue).

Preparation and loading of nanoparticles and microparticles are wellknown in the art.

As one example, liposomes can be prepared from dipalmitoylphosphatidylcholine (DPPC) or egg phosphatidylcholine (PC) because thislipid has a low heat transition. Liposomes are made using standardprocedures as known to one skilled in the art (e.g., Braun-Falco et al.,(Eds.), Griesbach Conference, Liposome Dermatics, Springer-Verlag,Berlin (1992), pp. 69 81; 91 117 which is expressly incorporated byreference herein). Polycaprolactone, poly(glycolic) acid, poly(lactic)acid, polyanhydride or lipids can be formulated as microspheres. As anillustrative example, the present compounds can be mixed with polyvinylalcohol (PVA), the mixture then dried and coated with ethylene vinylacetate, then cooled again with PVA. In a liposome, the presentcompounds can be within one or both lipid bilayers, in the aqueousbetween the bilayers, or within the center or core. Liposomes can bemodified with other molecules and lipids to form a cationic liposome.Liposomes can also be modified with lipids to render their surface morehydrophilic which increases their circulation time in the bloodstream.The thus-modified liposome has been termed a “stealth” liposome, or along-lived liposome, as described in U.S. Pat. No. 6,258,378, and inStealth Liposomes, Lasic and Martin (Eds.) 1995 CRC Press, London, whichare expressly incorporated by reference herein. Encapsulation methodsinclude detergent dialysis, freeze drying, film forming, injection, asknown to one skilled in the art and disclosed in, for example, U.S. Pat.No. 6,406,713 which is expressly incorporated by reference herein in itsentirety. Optionally, the present compositions and methods include amicelle delivery system, for example, involving one or more PEG-basedamphiphilic polymers developed for drug delivery including:PEG-poly(ε-caprolactone), PEG-poly(amino acid), PEG-polylactide orPEG-phospholipid constructs; a cross linked poly(acrylic acid) polymersystem, a phospholipid-based system and/or block copolymer systemscomprising one or more of the following polymer blocks: a poly(lacticacid) polymer block; a poly(propylene glycol) polymer block; apoly(amino acid) polymer block; a poly(ester) polymer block; apoly(ε-caprolactone) polymer block; a poly(ethylene glycol) block, apoly(acrylic acid) block; a polylactide block; a polyester block; apolyamide block; a polyanhydride block; a polyurethane block; apolyimine block; a polyurea block; a polyacetal block; a polysaccharideblock; and a polysiloxane block.

Every formulation or combination of components described or exemplifiedherein can be used to practice the invention, unless otherwise stated.

(i) Binding Agents

Binding agents include, but are not limited to, corn starch, potatostarch, or other starches, gelatin, natural and synthetic gums such asacacia, sodium alginate, alginic acid, other alginates, powderedtragacanth, guar gum, cellulose and its derivatives (e.g., ethylcellulose, cellulose acetate, carboxymethyl cellulose calcium, sodiumcarboxymethyl cellulose), polyvinyl pyrrolidone, methyl cellulose,pre-gelatinized starch, hydroxypropyl methyl cellulose, (e.g., Nos.2208, 2906, 2910), microcrystalline cellulose, and mixtures thereof.Suitable forms of microcrystalline cellulose include, for example, thematerials sold as AVICEL-PH-101, AVICEL-PH-103 and AVICEL-PH-105(available from FMC Corporation, American Viscose Division, AvicelSales, Marcus Hook, Pa., USA). An exemplary suitable binder is a mixtureof microcrystalline cellulose and sodium carboxymethyl cellulose sold asAVICEL RC-581 by FMC Corporation.

(ii) Fillers

Fillers include, but are not limited to, talc, calcium carbonate (e.g.,granules or powder), lactose, microcrystalline cellulose, powderedcellulose, dextrates, kaolin, mannitol, silicic acid, sorbitol, starch,pre-gelatinized starch, and mixtures thereof.

(iii) Lubricants

Lubricants include, but are not limited to, calcium stearate, magnesiumstearate, mineral oil, electromagnetic radiation mineral oil, glycerin,sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid,sodium lauryl sulfate, talc, hydrogenated vegetable oil (e.g., peanutoil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, andsoybean oil), zinc stearate, ethyl oleate, ethyl laurate, agar, andmixtures thereof. Additional lubricants include, for example, a syloidsilica gel (AEROSIL 200, manufactured by W.R. Grace Co. of Baltimore,Md., USA), a coagulated aerosol of synthetic silica (marketed by DeaussaCo. of Plano, Tex., USA), CAB-O-SIL (a pyrogenic silicon dioxide productsold by Cabot Co. of Boston, Mass., USA), and mixtures thereof.

(iv) Disintegrants

Disintegrants include, but are not limited to, agar-agar, alginic acid,calcium carbonate, microcrystalline cellulose, croscarmellose sodium,crospovidone, polacrilin potassium, sodium starch glycolate, potato ortapioca starch, other starches, pre-gelatinized starch, other starches,clays, other algins, other celluloses, gums, and mixtures thereof.

Tablets or capsules can optionally be coated by methods well known inthe art. If binders and/or fillers are used with a compound/bioconjugateof the invention, they are typically formulated as about 50 to about 99weight percent of the compound/bioconjugate. In one aspect, about 0.5 toabout 15 weight percent of disintegrant, and particularly about 1 toabout 5 weight percent of disintegrant, can be used in combination withthe compound. A lubricant can optionally be added, typically in anamount of less than about 1 weight percent of the compound/bioconjugate.Techniques and pharmaceutically acceptable additives for making solidoral dosage forms are described in Marshall, SOLID ORAL DOSAGE FORMS,Modern Pharmaceutics (Banker and Rhodes, Eds.), 7:359-427 (1979). Otherformulations are known in the art.

Liquid preparations for oral administration can take the form ofsolutions, syrups or suspensions. Alternatively, the liquid preparationscan be presented as a dry product for constitution with water or othersuitable vehicle before use. Such liquid preparations can be prepared byconventional means with pharmaceutically acceptable additives such assuspending agents (e.g., sorbitol syrup, cellulose derivatives orhydrogenated edible fats); emulsifying agents (e.g., lecithin oracacia); non-aqueous vehicles (e.g., almond oil, oily esters, ethylalcohol or fractionated vegetable oils); and/or preservatives (e.g.,methyl or propyl-p-hydroxybenzoates or sorbic acid). The preparationscan also contain buffer salts, flavoring, coloring, perfuming andsweetening agents as appropriate. Preparations for oral administrationcan also be formulated to achieve controlled release of thecompound/bioconjugate. Oral formulations preferably contain 10% to 95%compound/bioconjugate. In addition, a compound/bioconjugate of thepresent invention can be formulated for buccal administration in theform of tablets or lozenges formulated in a conventional manner. Othermethods of oral delivery of compounds/bioconjugates of the inventionwill be known to the skilled artisan and are within the scope of theinvention.

Formulation 1

Hard gelatin capsules are prepared using the following ingredients:

TABLE F1 Ingredients (mg/capsule) Active Ingredient 250.0 Starch 305.0Magnesium stearate 5.0

The above ingredients are mixed and filled into hard gelatin capsules in560 mg quantities.

Formulation 2

A tablet formula is prepared using the following ingredients:

TABLE F2 Ingredients (mg/tablet) Active Ingredient 250.0 Cellulose,microcrystalline 400.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0

The components are blended and compressed to form tablets, each weighing665 mg.

Formulation 3

A dry powder inhaler formulation is prepared containing the followingcomponents:

TABLE F3 Ingredients Weight % Active ingredient 5 Lactose 95

The active ingredient is mixed with the lactose and the mixture is addedto a dry powder inhaling appliance.

Formulation 4

Tablets, each containing 60 mg of active ingredient, are prepared asfollows:

TABLE F4 Ingredients milligrams Active ingredient 60.0 Starch 45.0Microcrystalline cellulose 35.0 Polyvinylpyrrolidone (as 10% solution inwater) 4.0 Sodium carboxymethyl starch 4.5 Magnesium stearate 0.5 Talc1.0 Total 150.0

The active ingredient, starch and cellulose are passed through a No. 20mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinylpyrrolidone is mixed with the resultant powders which are thenpassed through a 16 mesh U.S. sieve. The granules as produced are driedat 50-60° C. and passed through a 16 mesh U.S. sieve. The sodiumcarboxymethyl starch, magnesium stearate, and talc, previously passedthrough a No. 30 mesh U.S. sieve, are then added to the granules which,after mixing, are compressed on a tablet machine to yield tablets eachweighing 150 mg.

Formulation 5

Capsules, each containing 80 mg of active ingredient are made asfollows:

TABLE F5 Ingredients milligrams Active ingredient 80.0 Starch 109.0Magnesium stearate 1.0 Total 190.0

The active ingredient, cellulose, starch, and magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 190 mg quantities.

Formulation 6

Suppositories, each containing 225 mg of active ingredient, are made asfollows:

TABLE F6 Ingredients milligrams Active Ingredient 225 Saturated fattyacid glycerides to 2000

The active ingredient is passed through a No. 60 mesh U.S. sieve andsuspended in the saturated fatty acid glycerides previously melted usingthe minimum heat necessary. The mixture is then poured into asuppository mold of nominal 2.0 g capacity and allowed to cool.

Formulation 7

Suspensions, each containing 50 mg of active ingredient per 5.0 ml doseare made as follows:

TABLE F7 Ingredients milligrams Active ingredient 50.0 mg Xanthan gum4.0 mg Sodium carboxymethyl cellulose (11%) Microcrystalline cellulose(89%) 50.0 mg Sucrose 1.75 g Sodium benzoate 10.0 mg Flavor q.v. Colorq.v. Purified water to 5.0 ml

The active ingredient, sucrose and xantham gum are blended, passedthrough a No. 10 mesh U.S. sieve, and mixed with a previously madesolution of the microcrystalline cellulose and sodium carboxymethylcellulose in water. The sodium benzoate, flavor, and color are dilutedwith some of the water and added with stirring. Sufficient water is thenadded to produce the required volume.

Formulation 8

Capsules, each containing 150 mg of active ingredient, are made asfollows:

TABLE F8 Ingredients milligrams Active ingredient 150.0 Starch 407.0Magnesium stearate 3.0 Total 560.0

The active ingredient, cellulose, starch, and magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 560 mg quantities.

In the formulation examples, the “active ingredient” is a compound ofthe invention.

Kits

Various embodiments of the present invention include kits. Such kits caninclude a compound/bioconjugate of the present invention, optionally oneor more ingredients for preparing a pharmaceutically acceptableformulation of the compound/bioconjugate, and instructions for use(e.g., administration). When supplied as a kit, different components ofa compound/bioconjugate formulation can be packaged in separatecontainers and admixed immediately before use. Such packaging of thecomponents separately can, if desired, be presented in a pack ordispenser device which can contain one or more unit dosage formscontaining the compound/bioconjugate. The pack can, for example,comprise metal or plastic foil such as a blister pack. Such packaging ofthe components separately can also, in certain instances, permitlong-term storage without losing activity of the components. Inaddition, if more than one route of administration is intended or morethan one schedule for administration is intended, the differentcomponents can be packaged separately and not mixed prior to use. Invarious embodiments, the different components can be packaged in onecombination for administration together.

It is further contemplated that the compounds and salts of thisinvention can be used in the form of a kit that is suitable for use inperforming the methods described herein, packaged in a container. Thekit can contain the compound or compounds and, optionally, appropriatediluents, devices or device components suitable for administration andinstructions for use in accordance with the methods of the invention.The devices can include parenteral injection devices, such as syringesor transdermal patch or the like. Device components can includecartridges for use in injection devices and the like. In one aspect, thekit includes a first dosage form including a compound or salt of thisinvention and a second dosage form including another active ingredientin quantities sufficient to carry out the methods of the invention. Thefirst dosage form and the second dosage form together can include atherapeutically effective amount of the compounds for treating thetargeted condition(s).

In certain embodiments, kits can be supplied with instructionalmaterials. Instructions can be printed on paper or other substrate,and/or can be supplied as an electronic-readable medium, such as afloppy disc, mini-CD-ROM, CD-ROM, DVD-ROM, Zip disc, videotape, audiotape, and the like. In an embodiment, detailed instructions are notphysically associated with the kit; instead, a user can be directed toan Internet web site specified by the manufacturer or distributor of thekit, or supplied as electronic mail, for example.

Kits can include reagents in separate containers such as, for example,sterile water or saline to be added to a lyophilized active componentpackaged separately. For example, sealed glass ampules can containlyophilized superoxide dismutase mimetics and in a separate ampule,sterile water, sterile saline or sterile each of which has been packagedunder a neutral non-reacting gas, such as nitrogen. Ampules can consistof any suitable material, such as glass, organic polymers, such aspolycarbonate, polystyrene, ceramic, metal or any other materialtypically employed to hold reagents. Other examples of suitablecontainers include bottles that can be fabricated from similarsubstances as ampules, and envelopes that can consist of foil-linedinteriors, such as aluminum or an alloy. Other containers include testtubes, vials, flasks, bottles, syringes, and the like. Containers canhave a sterile access port, such as a bottle having a stopper that canbe pierced by a hypodermic injection needle. Other containers can havetwo compartments that are separated by a readily removable membrane thatupon removal permits the components to mix. Removable membranes can beglass, plastic, rubber, and the like.

Example 6: Synthesis

The compounds of the invention can be synthesized according to themethods in references known to the art and using the provided proceduresand modifications thereof known to one of ordinary skill in the art.Exemplary synthetic methods is provided in references including: M. E.Vol'pin, Russian Chemical Reviews, Vol. 29, No. 3 (1960), 129-160;Cowles, JACS, Vol. 79 (1957), 1093-1095; Hess, Tetrahedron, Vol. 31(1975), 295-298; Anderson, J. Org. Chem., Vol. 29 (1964), 1373-1377;Nozoe, Chemistry and Industry (1954), 1357-1358; U.S. Pat. No.5,846,730; Shevyakov, J. Phys. Chem., Vol. 107 (2003), 3295-3299;Murata, Chem. Phys. Lett., Vol. 13 (1972), 101-104; Pham, Angew. Chem.Int. Ed., Vol. 41 (2002) 3659-3662; Xu, Bioorganic & Medicinal ChemistryLetters, Vol. 11 (2001) 2045-2047; Wu, Anti-cancer drug design, Vol. 15(2000), 287-293.

Exemplary synthetic schemes illustrating synthesis of severalembodiments of the invention are shown below:

Peptide coupling can be carried out as known in the art by reacting a—COOH or —CONH(CH₂)_(a)CO₂H functional group on the azulene or azulenederivative using the reagent H₂N-Peptide and Coupling Agents, includingbut not limited to:DCCEDCDCC, NHSEDC, NHSEDC, HOBtPyBOPPyBrOPHATUHBTUAlso provided below are exemplary synthetic methods for embodiments ofthe invention:Solution Preparation of Preparation of Azulene-Peptide Conjugate 16where Peptide=SFFYLRS (SEQ ID NO:1)

A mixture of 1-azulenecarboxylic acid (1 mmol), 1-hydroxybenzotriazole(1.2 mmol), and triethylamine (1.2 mmol) in DMF is stirred at ambienttemperature for about 30 minutes. Thereafter, EDC (1.2 mmol) is thenadded and the entire mixture is stirred at ambient temperature for about16 hours. The reaction mixture is then poured onto ethyl acetate toprecipitate the product. Excess solvent is decanted off and the residueis repeatedly washed with ethyl acetate to remove excess DMF and thecrude material is purified by HPLC.

Automated Procedure for Preparation of Azulene-Peptide Conjugate 16where Peptide=SFFYLRS (SEQ ID NO:1)

A typical procedure for the preparation of azulene-peptide conjugate 16using an automated peptide synthesizer is described. It should be notedthat other azulene fluorophores may be conjugated to leukemia bindingand other targeting groups by the same procedure. The leukemia cellbinding peptide conjugate pyrazine-S-F-F-Y-L-R-S(SEQ ID NO:1) (16) wasprepared by fluorenylmethyloxycarbonyl (Fmoc) solid phase peptidesynthesis strategy with a commercial automated peptide synthesizer. Thefirst peptide cartridge contained Wang resin pre-loaded with an amideresin on 25 μmole scale. The amino acid cartridges each containing S, F,F, Y, L, R, and S amino acids were placed on the peptide synthesizer andthe product was synthesized from the C- to the N-terminal sequence.Coupling of the Fmoc-protected amino acids to the resin-bound freeterminal amine was carried out with2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate(HBTU, 75 μmol)/N-hydroxybenzotriazole (HOBt, 75 μmol). Each Fmocprotecting group on solid support was removed with 20% piperidine indimethylformamide before the subsequent amino acid was coupled to it.The last cartridge contained the 1-azulenecarboxylic acid which wascoupled to the

peptide automatically, thus avoiding the need for post-syntheticmanipulations. After the synthesis, the product was cleaved from thesolid support with a cleavage mixture containing trifluoroacetic acid(85%):water (5%):phenol (5%):thioanisole (5%) for 6 hours. The peptideconjugate was precipitated with t-butyl methyl ether, collected byfiltration, and purified by HPLC to give 60 mg of the desiredpyrazine-peptide conjugate as a magenta powder.

Table of Sequences SEQ ID NO: SEQUENCE 1 S-F-F-Y-L-R-SPreparation of Azulene-mPEG₁₂ Derivative

A mixture of the 1-azulene carboxylic acid (172 mg, 1.0 mmol) andm-PEG₁₂-amine (560 mg, 1.0 mmol) in anhydrous methylene chloride (10 mL)was stirred at ambient temperature for 15 minutes. Thereafter,1-hydroxybenzotriazole (168 mg, 1.1 mmol) and ethyl dimethylaminopropylcarbodiimide (EDC) hydrochloride (210 mg 1.1 mmol) was added, and theentire mixture was stirred at ambient temperature for 16 hours. Thereaction mixture was treated with 0.1 M HCl (10 mL) and the organiclayer was separated, washed with water, dried over anhydrous sodiumsulfate, filtered, and the filtrate evaporated to dryness in vacuo. Thecrude product was purified by flash chromatography usingchloroform-methanol gradient (0 to 5% over 45 minutes) to give thedesired material as magenta gum. LR/MS, 714.7 (M+H⁺). UV, λ_(max), 548nm.

Statements Regarding Incorporation by Reference and Variations

All references throughout this application, for example patent documentsincluding issued or granted patents or equivalents; patent applicationpublications; and non-patent literature documents or other sourcematerial; are hereby incorporated by reference herein in theirentireties, as though individually incorporated by reference, to theextent each reference is at least partially not inconsistent with thedisclosure in this application (for example, a reference that ispartially inconsistent is incorporated by reference except for thepartially inconsistent portion of the reference).

The terms and expressions which have been employed herein are used asterms of description and not of limitation, and there is no intention inthe use of such terms and expressions of excluding any equivalents ofthe features shown and described or portions thereof, but it isrecognized that various modifications are possible within the scope ofthe invention claimed. Thus, it should be understood that although theinvention has been specifically disclosed by preferred embodiments,exemplary embodiments and optional features, modification and variationof the concepts herein disclosed may be resorted to by those skilled inthe art, and that such modifications and variations are considered to bewithin the scope of this invention as defined by the appended claims.The specific embodiments provided herein are examples of usefulembodiments of the invention and it will be apparent to one skilled inthe art that the invention may be carried out using a large number ofvariations of the devices, device components, methods steps set forth inthe present description. As will be obvious to one of skill in the art,methods and devices useful for the present methods can include a largenumber of optional composition and processing elements and steps.

When a group of substituents is disclosed herein, it is understood thatall individual members of that group and all subgroups, including anyisomers, enantiomers, and diastereomers of the group members, aredisclosed separately. When a Markush group or other grouping is usedherein, all individual members of the group and all combinations andsubcombinations possible of the group are intended to be individuallyincluded in the disclosure. When a compound is described herein suchthat a particular isomer, enantiomer or diastereomer of the compound isnot specified, for example, in a formula or in a chemical name, thatdescription is intended to include each isomers and enantiomer of thecompound described individual or in any combination. Additionally,unless otherwise specified, all isotopic variants of compounds disclosedherein are intended to be encompassed by the disclosure. For example, itwill be understood that any one or more hydrogens in a moleculedisclosed can be replaced with deuterium or tritium. Isotopic variantsof a molecule are generally useful as standards in assays for themolecule and in chemical and biological research related to the moleculeor its use. Methods for making such isotopic variants are known in theart. Specific names of compounds are intended to be exemplary, as it isknown that one of ordinary skill in the art can name the same compoundsdifferently.

Many of the molecules disclosed herein contain one or more ionizablegroups [groups from which a proton can be removed (e.g., —COOH) or added(e.g., amines) or which can be quaternized (e.g., amines)]. All possibleionic forms of such molecules and salts thereof are intended to beincluded individually in the disclosure herein. With regard to salts ofthe compounds herein, one of ordinary skill in the art can select fromamong a wide variety of available counterions those that are appropriatefor preparation of salts of this invention for a given application. Inspecific applications, the selection of a given anion or cation forpreparation of a salt may result in increased or decreased solubility ofthat salt.

Every formulation or combination of components described or exemplifiedherein can be used to practice the invention, unless otherwise stated.

The present compositions, preparations and formulations can be used bothas a diagnostic agent as well as a photodynamic therapeutic agentconcomitantly. For example, an effective amount of the presentcompositions, preparations and formulations in a pharmaceuticallyacceptable formulation is administered to a patient. Administration isfollowed by a procedure that combines photodiagnosis and phototherapy.For example, a composition comprising compounds for combinedphotodiagnosis and phototherapy is administered to a patient and itsconcentration, localization, or other parameters is determined at thetarget site of interest. More than one measurement may be taken todetermine the location of the target site. The time it takes for thecompound to accumulate at the target site depends upon factors such aspharmcokinetics, and may range from about thirty minutes to two days.Once the site is identified, the phototherapeutic part of the proceduremay be done either immediately after determining the site or before theagent is cleared from the site. Clearance depends upon factors such aspharmacokinetics.

The present compositions, preparations and formulations can beformulated into diagnostic or therapeutic compositions for enteral,parenteral, topical, aerosol, inhalation, or cutaneous administration.Topical or cutaneous delivery of the compositions, preparations andformulations may also include aerosol formulation, creams, gels,solutions, etc. The present compositions, preparations and formulationsare administered in doses effective to achieve the desired diagnosticand/or therapeutic effect. Such doses may vary widely depending upon theparticular compositions employed in the composition, the organs ortissues to be examined, the equipment employed in the clinicalprocedure, the efficacy of the treatment achieved, and the like. Thesecompositions, preparations and formulations contain an effective amountof the composition(s), along with conventional pharmaceutical carriersand excipients appropriate for the type of administration contemplated.These compositions, preparations and formulations may also optionallyinclude stabilizing agents and skin penetration enhancing agents.

Methods of this invention comprise the step of administering an“effective amount” of the present diagnostic and therapeuticcompositions, formulations and preparations containing the presentcompounds, to diagnose, image, monitor, evaluate, reduce or regulate abiological condition and/or disease state in a patient. The term“effective amount,” as used herein, refers to the amount of thediagnostic and therapeutic formulation, that, when administered to theindividual is effective diagnosis, image, monitor, evaluate treat,reduce or regulate a biological condition and/or disease state. As isunderstood in the art, the effective amount of a given composition orformulation will depend at least in part upon, the mode ofadministration (e.g. intravenous, oral, topical administration), anycarrier or vehicle employed, and the specific individual to whom theformulation is to be administered (age, weight, condition, sex, etc.).The dosage requirements need to achieve the “effective amount” vary withthe particular formulations employed, the route of administration, andclinical objectives. Based on the results obtained in standardpharmacological test procedures, projected daily dosages of activecompound can be determined as is understood in the art. As used herein,“treat” means reduce or regulate a biological condition and/or diseasestate in a patient.

Any suitable form of administration can be employed in connection withthe diagnostic and therapeutic formulations of the present invention.The diagnostic and therapeutic formulations of this invention can beadministered intravenously, in oral dosage forms, intraperitoneally,subcutaneously, or intramuscularly, all using dosage forms well known tothose of ordinary skill in the pharmaceutical arts.

The diagnostic and therapeutic formulations of this invention can beadministered alone, but may be administered with a pharmaceuticalcarrier selected upon the basis of the chosen route of administrationand standard pharmaceutical practice.

The diagnostic and therapeutic formulations of this invention andmedicaments of this invention may further comprise one or morepharmaceutically acceptable carrier, excipient, buffer, emulsifier,surfactant, electrolyte or diluent. Such compositions and medicamentsare prepared in accordance with acceptable pharmaceutical procedures,such as, for example, those described in Remingtons PharmaceuticalSciences, 17th edition, ed. Alfonoso R. Gennaro, Mack PublishingCompany, Easton, Pa. (1985).

Whenever a range is given in the specification, for example, atemperature range, a time range, or a composition or concentrationrange, all intermediate ranges and subranges, as well as all individualvalues included in the ranges given are intended to be included in thedisclosure. It will be understood that any subranges or individualvalues in a range or subrange that are included in the descriptionherein can be excluded from the claims herein. As used herein, rangesspecifically include the values provided as endpoint values of therange. As used herein, ranges specifically include all the integervalues of the range. For example, a range of 1 to 100 specificallyincludes the end point values of 1 and 100.

All patents and publications mentioned in the specification areindicative of the levels of skill of those skilled in the art to whichthe invention pertains.

As used herein, “comprising” is synonymous with “including,”“containing,” or “characterized by,” and is inclusive or open-ended anddoes not exclude additional, unrecited elements or method steps. As usedherein, “consisting of” excludes any element, step, or ingredient notspecified in the claim element. As used herein, “consisting essentiallyof” does not exclude materials or steps that do not materially affectthe basic and novel characteristics of the claim. In each instanceherein any of the terms “comprising”, “consisting essentially of” and“consisting of” may be replaced with either of the other two terms. Theinvention illustratively described herein suitably may be practiced inthe absence of any element or elements, limitation or limitations whichis not specifically disclosed herein. Any claim that is written inmultiple dependent form or is a multiple dependent claim dependent onother multiple dependent claims is intended to include all claimedsubject matter to the extent that each aspect can be included in aseparate claim if necessary or desired.

One of ordinary skill in the art will appreciate that startingmaterials, biological materials, reagents, synthetic methods,purification methods, analytical methods, assay methods, and biologicalmethods other than those specifically exemplified can be employed in thepractice of the invention without resort to undue experimentation. Allart-known functional equivalents, of any such materials and methods areintended to be included in this invention. The terms and expressionswhich have been employed are used as terms of description and not oflimitation, and there is no intention in the use of such terms andexpressions of excluding any equivalents of the features shown anddescribed or portions thereof, but it is recognized that variousmodifications are possible within the scope of the invention claimed.Thus, it should be understood that although the invention has beenspecifically disclosed by preferred embodiments and optional features,modification and variation of the concepts herein disclosed may beresorted to by those skilled in the art, and that such modifications andvariations are considered to be within the scope of this invention asdefined by the appended claims.

The disclosures of the publications listed herein including thepublications listed below are herein incorporated by reference in theirentireties.

REFERENCES

-   Shevyakov, V. et al. Orbital control of the color and excited state    properties of formylated and fluorinated derivatives of azulene. J.    Phys. Chem. 2003, 107, 3295-3299.-   Murata, S. et al. Fluorescence yields of azulene derivatives. Chem.    Phys. Lett. 1972, 13(2), 101-104.-   Hess, B. A. Jr. et al. The aromaticity of heterocycles containing    the imine nitrogen. Tetrahedron 1975, 31, 295-298.-   Cowles, E. J. The effects of substituents at the 1- and 3-positions    of the visible absorption spectrum of azulene. J. Am. Chem. Soc.    1957, 79, 1093-1095.-   Anderson, A. G. et al. Some new reactions and derivatives of    azulene. J. Org. Chem. 1964, 29, 1373-1377.-   Nozoe, T. et al. Synthesis of 1-azaazulene and its derivatives.    Chem. Ind. (London, UK) 1954, 1357-1358.

What is claimed is:
 1. A compound of the formula (FX10):

wherein: each of L² to L⁸ is independently selected from C₁-C₁₀alkylene, C₃-C₁₀ cycloalkylene, C₂-C₁₀ alkenylene, C₃-C₁₀cycloalkenylene, C₂-C₁₀ alkynylene, ethenylene, ethynylene, phenylene,1-aza-2,5-dioxocyclopentylene, (CH₂CH₂O)_(m)—, —(CHOH)_(m)—, or1,4-diazacyclohexylene; each m is independently selected from an integerselected from the range of 1 to 100; each q is independently selectedfrom 0 or 1; each of W² to W⁸ is independently selected from a singlebond, —(CH₂)_(n)—, —O(CH₂)_(n)—, —(CH₂)_(n)O—, —(HCCH)_(n)—, —O—, —S—,—SO—, —SO₂—, —SO₃—, —OSO₂—, —NR⁹—, —CO—, —COO—, —OCO—, —OCOO—, —CONR¹⁰—,—NR¹¹CO—, —OCONR¹²—, —NR¹³COO—, —NR¹⁴CONR¹⁵—, —NR¹⁶CSNR¹⁷, —O(CH₂)_(n)—,—NR¹⁸(CH₂)_(n)—, —CO(CH₂)_(n)—, —COO(CH₂)_(n)—, —OCO(CH₂)_(n)—,—OCOO(CH₂)_(n)—, —CONR¹⁹(CH₂)_(n)—, —CONR²⁰(CH₂)_(n)(OCH₂CH₂)_(u)—,—NR²¹CO(CH₂)_(n)—, —OCONR²²(CH₂)_(n)—, —NR²³COO(CH₂)_(n)—,—NR²⁴CONR²⁵(CH₂)_(n)—, —NR²⁶CSNR²⁷(CH₂)_(n)—,—O(CH₂)_(n)NR²⁸CO(CH₂)_(n)—,—CO(CH₂)_(n)(CH₂OCH₂)_(n)(CH₂)_(n)NR²⁹(CH₂)_(n)NR³⁰CO—, —NR⁶⁹SR⁷⁰—, or—CO(CH₂)_(n)NR³¹CO—; each n is independently selected from an integerselected from the range of 1 to 10; each of R⁹ to R³¹ and each of R⁶⁹ toR⁷⁰ is independently selected from hydrogen, C₁-C₂₀ alkyl, or C₅-C₃₀aryl; each R³ to R⁸ is independently selected from hydrogen, C₁-C₂₀alkyl, C₅-C₃₀ aryl, C₃-C₂₀ acyl, C₂-C₂₀ alkenyl, C₂-C₂₀ alkynyl, C₅-C₂₀alkylaryl, C₁-C₆ alkoxycarbonyl, halo, halomethyl, dihalomethyl,trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷—,—SO₂R³⁸, —SO₂OR³⁹, —SO₂NR⁴⁰R⁴¹, —PO₃R⁴²R⁴³, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷,—NR⁴⁸COR⁴⁹, —CH₂(CHOH)_(n)R⁵⁰, —(CH₂CH₂O)_(n)R⁵¹, —CH(R⁵²)CO₂H,—CH(R⁵³)NH₂, TG¹ to TG⁸, PS¹ to PS⁸, or FL¹ to FL⁸; R² is TG¹, PS¹ orFL¹; each u is independently selected from an integer selected from therange of 1 to 25; each of R³² to R⁵⁵ is independently selected fromhydrogen or C₁-C₁₀ alkyl; each of TG¹ to TG⁸ is independently selectedfrom an amino acid, a peptide, a protein, a nucleoside, a nucleotide, anenzyme, a carbohydrate, a glycomimetic, an oligomer, a lipid, a polymer,an antibody, an antibody fragment, a mono- or polysaccharide comprising1 to 50 carbohydrate units, a glycopeptide, a glycoprotein, apeptidomimetic, a drug, a drug mimic, a hormone, a receptor, a metalchelating agent, a radioactive or nonradioactive metal complex, a mono-or polynucleotide comprising 1 to 50 nucleic acid units, a polypeptidecomprising 2 to 30 amino acid units, or an echogenic agent; each of PS¹to PS⁸ is independently selected from at least one azide, azo, diazo,oxaza, diaza, dithia, thioxa, dioxa, phthalocyanine, rhodamine, andporphyrin, and wherein each of PS¹ to PS⁸ is capable of reacting toproduce one or more free radicals selected from nitrenes, carbenes, andsinglet oxygen; and each of FL¹ to FL⁸ is independently selected from afluorescent group selected from the group consisting of anaphthoquinone, an anthracene, an anthraquinone, a phenanthrene, atetracene, a naphthacenedione, a pyridine, a quinoline, an isoquinoline,an indole, an isoindole, a pyrrole, an imidiazole, a pyrazole, apyrazine, a purine, a benzimidazole, a benzofuran, a dibenzofuran, acarbazole, an acridine, an acridone, a phenanthridine, a thiophene, abenzothiophene, a dibenzothiophene, a xanthene, a xanthone, a flavone, acoumarin, a phenoxazine, a phenothiazine, a phenoselenazine, a cyanine,an indocyanine, and an azo compound; wherein any adjacent R³ to R⁸optionally combines with one or two —CR⁵⁴R⁵⁵ groups, to form C₃-C₇cycloalkyl, C₃-C₇ heterocycloalkyl, C₆ aryl, or C₅-C₆ heteroaryl;wherein at least one of R³ to R⁸ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵,—NR⁴⁶R⁴⁷, or NR⁴⁸COR⁴⁹; and wherein at least one of R³ to R⁸ is halo,trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —OSR³⁷,—SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl; or wherein at least one of-(L²)_(q)-W²—R², (L³)_(q)-W³—R³, (L⁴)_(q)-W⁴—R⁴, (L⁵)_(q)-W⁵—R⁵,(L⁶)_(q)-W⁶—R⁶, -(L⁷)_(q)-W⁷—R⁷, or -(L⁸)_(q)-W⁸—R⁸ includes—(OCH₂CH₂)_(u)—.
 2. The compound of claim 1, wherein at least one of R³to R⁸ is independently selected from C₁-C₆ alkyl, —OR⁴⁴, —SR⁴⁵,—NR⁴⁶R⁴⁷, or NR⁴⁸COR⁴⁹.
 3. The compound of claim 1, wherein at least oneof R³ to R⁸ is independently selected from halo, trihalomethyl, —CN,—CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, or —SO₂NR⁴⁰R⁴¹. 4.The compound of claim 1, having R group substituent pairings (R³ andR⁴); (R⁴ and R⁵); (R⁵ and R⁶); (R⁶ and R⁷); (R⁷ and R⁸); (R³ and R⁵); or(R⁴ and R⁷), wherein one of the identified R groups in the substituentpairings is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or NR⁴⁸COR⁴⁹ and theother of the identified R groups in the substituent pairings is halo,trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸,or —SO₂NR⁴⁰R⁴¹.
 5. The compound of claim 1, wherein at least one of R³to R⁸ is FL¹, FL², FL³, FL⁴, FL⁵, FL⁶, FL⁷ or FL⁸, or R² is FL¹.
 6. Thecompound of claim 1, wherein each of FL¹ to FL⁸ is independentlyselected from a naphthoquinone, an anthraquinone, a naphthacenedione, apyrazine, an acridine, an acridone, a phenanthridine, adibenzothiophene, a xanthene, a xanthone, a flavone, a coumarin, aphenoxazine, a phenothiazine, a phenoselenazine, a cyanine, anindocyanine, or an azo compound.
 7. The compound of claim 1, wherein atleast one of R³ to R⁸ is PS¹, PS², PS³, PS⁴, PS⁵, PS⁶, PS⁷ or PS⁸, or R²is PS¹.
 8. The compound of claim 1, wherein at least one of PS¹ to PS⁸is independently selected from an azide, an azo, a diazo, an oxaza, adiaza, a thioxa, a phthalocyanine, a rhodamine, or a porphyrin group. 9.The compound of claim 1, wherein at least one of R³ to R⁸ is TG¹, TG²,TG³, TG⁴, TG⁵, TG⁶, TG⁷ or TG⁸, or R² is TG¹.
 10. The compound of claim1, wherein each of TG¹ to TG⁸ is independently selected from an aminoacid, a peptide, a protein, a nucleoside, a nucleotide, an enzyme, anantibody, an antibody fragment, a mono- or polysaccharide comprising 1to 50 carbohydrate units, a glycopeptide, a peptidomimetic, a drug, adrug mimic, or a hormone.
 11. The compound of claim 1, wherein: (b) oneof R³ and R⁴ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸CO⁴⁹, andthe other of R³ and R⁴ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹ or C₃-C₁₀ acyl; (c) one of R⁴and R⁵ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸CO⁴⁹, and theother of R⁴ and R⁵ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹ or C₃-C₁₀ acyl; (d) one of R⁵and R⁶ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹, and theother of R⁵ and R⁶ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl; (e) one ofR⁶ and R⁷ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹, andthe other of R⁶ and R⁷ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl; or (f) oneof R⁷ and R⁸ is C₁-C₁₀ alkyl, —OR⁴⁴, —SR⁴⁵, NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹, andthe other of R⁷ and R⁸ is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴,—COR³⁵, —NO₂, —SOR³⁶, —SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl.
 12. Thecompound of claim 1, wherein: (d) one of R³ and R⁵ is C₁-C₁₀ alkyl,—OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹, and the other of R³ and R⁵ ishalo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶,—SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl; or (e) one of R⁴ and R⁷ is C₁-C₁₀alkyl, —OR⁴⁴, —SR⁴⁵, —NR⁴⁶R⁴⁷, or —NR⁴⁸COR⁴⁹, and the other of R⁴ and R⁷is halo, trihalomethyl, —CN, —CO₂R³², —CONR³³R³⁴, —COR³⁵, —NO₂, —SOR³⁶,—SO₂R³⁸, —SO₂NR⁴⁰R⁴¹, or C₃-C₁₀ acyl.
 13. A pharmaceutical compositioncomprising: a compound of claim 1; and a pharmaceutically acceptableexcipient.